\r\n\tA number of advanced combustion technologies have been introduced to improve performance, fuel economy and emissions levels. Research in combustion technology has highlighted the importance of new fuels in reducing the petroleum dependence and achieving high efficiency with low pollutant formation. \r\n\tThe purpose of this book is to collect interesting and original studies on combustion methods, advanced combustion strategies and new fuels able to achieve efficiency improvements and environment compliance. \r\n\tContributions in which experimental, theoretical and computation approaches are applied to explore how fuel properties and composition affect advanced combustion systems and how advanced combustion technology can maximize engine efficiency and be environment-friendly are invited and appreciated.
",isbn:null,printIsbn:"979-953-307-X-X",pdfIsbn:null,doi:null,price:0,priceEur:0,priceUsd:0,slug:null,numberOfPages:0,isOpenForSubmission:!1,hash:"025ed42cace07d115f7220a5a7b26f56",bookSignature:"Prof. Ornella Chiavola",publishedDate:null,coverURL:"https://cdn.intechopen.com/books/images_new/8231.jpg",keywords:"Combustion process, Performance, Fuel consumption, Emissions levels, Efficiency improvements, Environmental impact, Sustainable fuels, Fossil fuels, New combustion methods, Combustion analysis, Pollutant formation, Efficiency",numberOfDownloads:null,numberOfWosCitations:0,numberOfCrossrefCitations:0,numberOfDimensionsCitations:0,numberOfTotalCitations:0,isAvailableForWebshopOrdering:!0,dateEndFirstStepPublish:"May 14th 2019",dateEndSecondStepPublish:"September 17th 2019",dateEndThirdStepPublish:"November 16th 2019",dateEndFourthStepPublish:"February 4th 2020",dateEndFifthStepPublish:"April 4th 2020",remainingDaysToSecondStep:"a year",secondStepPassed:!0,currentStepOfPublishingProcess:5,editedByType:null,kuFlag:!1,biosketch:null,coeditorOneBiosketch:null,coeditorTwoBiosketch:null,coeditorThreeBiosketch:null,coeditorFourBiosketch:null,coeditorFiveBiosketch:null,editors:[{id:"201229",title:"Prof.",name:"Ornella",middleName:null,surname:"Chiavola",slug:"ornella-chiavola",fullName:"Ornella Chiavola",profilePictureURL:"https://mts.intechopen.com/storage/users/201229/images/system/201229.jfif",biography:"Ornella Chiavola completed a PhD in Mechanical Engineering at Engineering Department of the University of ‘La Sapienza’ – Rome, Italy in 1999. \r\n\r\nShe is currently working as an Associate Professor at ‘ROMA TRE’ University, Engineering Department, where she teaches ‘Interaction between machines and environment’ at Master Degree of Mechanical Engineering and at Master Degree of Aeronautical Engineering. \r\n\r\nFrom 2012, she is a member of the Teaching Board of Mechanical and Industrial Engineering PhD at ‘ROMA TRE’ University. \r\nShe has published numerous articles in international journals and is a member of the editorial board for several journals concerning her field of research.",institutionString:"Roma Tre University",position:null,outsideEditionCount:0,totalCites:0,totalAuthoredChapters:"2",totalChapterViews:"0",totalEditedBooks:"0",institution:{name:"Roma Tre University",institutionURL:null,country:{name:"Italy"}}}],coeditorOne:null,coeditorTwo:null,coeditorThree:null,coeditorFour:null,coeditorFive:null,topics:[{id:"11",title:"Engineering",slug:"engineering"}],chapters:null,productType:{id:"1",title:"Edited Volume",chapterContentType:"chapter",authoredCaption:"Edited by"},personalPublishingAssistant:{id:"301331",firstName:"Mia",lastName:"Vulovic",middleName:null,title:"Mrs.",imageUrl:"https://mts.intechopen.com/storage/users/301331/images/8498_n.jpg",email:"mia.v@intechopen.com",biography:"As an Author Service Manager, my responsibilities include monitoring and facilitating all publishing activities for authors and editors. From chapter submission and review to approval and revision, copyediting and design, until final publication, I work closely with authors and editors to ensure a simple and easy publishing process. I maintain constant and effective communication with authors, editors and reviewers, which allows for a level of personal support that enables contributors to fully commit and concentrate on the chapters they are writing, editing, or reviewing. I assist authors in the preparation of their full chapter submissions and track important deadlines and ensure they are met. I help to coordinate internal processes such as linguistic review, and monitor the technical aspects of the process. As an ASM I am also involved in the acquisition of editors. 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1. Introduction
Neuropeptide- glutamic acid isoleucine (NEI) is a peptide related to reproduction. Although it also has an important function on behavior [2-6], we will focus here on the relationship between NEI and LH.
NEI is derived from the precursor pre- prohormone named pp melanin-concentrating hormone (pp-MCH). This precursor also gives rise to melanin concentrating hormone (MCH) and to neuropeptide-glycine-glutamic acid (NGE) [7-9].
Some studies have suggested a role of NGE at the level of the hypothalamus. For instance, NGE increases the number of neurofilaments and the production of synaptophysin in rat neurons within 18 days of development [10-12].
Immunoreactivity and mRNA expression of both MCH and NEI have been observed in certain regions of the central nervous system. The first region is the diencephalon including the rostralmedial part of the zona incerta, later refer to as the incerto-hypothalamic area (Ihy) by Sita et al. [13]; the three subdivisions (anterior, tuberal and posterior) of the lateral hypothalamus (LHA); the area between the dorsomedial and ventromedial nuclei of the hypothalamus, which Swanson [14] designated as the internuclear area, the anterior periventricular nucleus; and the dorso medial aspects of the tuberomammillary complex. The second region includes the olfatory tubercle, located in the basal forebrain. The third region includes the paramedian pontine reticular formation in the pons [7]. It is important to note that the highest concentration of MCH/NEI ir cells is found in the Ihy and the LHA. We found nearly all the cells in these two regions were immureactive for both MCH and NEI (mean +-S.E.M, 96 +-3%) (Fig.1 and 2)
Figure 1.
Diencephalic distribution of NEI-immunoreactivity-containing cells. Brightfield photomicrographs of immunoperoxidase material stained for NEI-ir. (A) NEI-ir-containing cells in the incerto-hypothalamic area. (B) NEI-ircontaining cells in the lateral hypothalamic area. Abbreviations: mt, mammilothalamic tract; bv, blood vessels; IHy, incerto hypothalamic area; 3v, third ventricle; ic, internal capsule; LHA, lateral hypothalamic area; f, fornix. Bar = 200 mm. - Reproduced from Bittencourt and Celis, 2008, with permission from Peptides (29:1441-50).
Figure 2.
Deduced structure of rat prepro-MCH. The relative positions of the amino (NH2) terminal signal peptide (SP), and the putative MCH, NEI and NGE sequences at the carboxy (COOH) terminus are indicated. The amino acids sequence of NEI is expanded below it. The putative proteolitic processing sites are marked with arrowheads. - Reproduced from Bittencourt and Celis, 2008, with permission from Peptides (29:1441-50).
Peptides such as NGE, NEI and MCH are highly conserved among vertebrates, being abundant and widely distributed in the brain, suggesting that they could be performing important physiological functions. NEI is a 13 aminoacid peptide and has an extensive distribution in the central nervous system (CNS), acting as a neurotransmitter or neuromodulator [15-17]. Neurotranmitter NEI induces grooming behavior, locomotor activity, and stimulates sexual receptivity in female rats [1-6]. NEI injections can modify the levels of noradrenaline and dopamine in specific areas of the brain [4], and earlier studies demonstrated that MCH injection into the preoptic area or median eminence induces luteinizing hormone (LH) secretion. Subsequent studies have shown that modified circulating hormonal levels might modulate ppMCH neurons [18] It has been reported that treatment with 17β-estradiol increases MCH and NEI immunoreactivity throughout the entire diencephalon of ovariectomized cynomologous monkeys [18]. Similar effects were observed in ovariectomized rats receiving no gonadal estradiol treatment [19]. Levels of ppMCH mRNA are only increased in the medial zone incerta (ZI) [15], which houses the A13 group, a collection of dopaminergic neurons that have been previously demonstrated to play a stimulating role in gonadotropin release [20].
More recently, various studies have explored the potential mechanisms by which MCH induces LH secretion [21-27]. Surprisingly, few have examined the role played by NEI in this process.
2. Ovarian steroids effects
Viale et al (1999) proposed a central role of the MCH/NEI neuronal system in the regulation of reproductive functions in rats [18]. In fact, the MCH/NEI system of immunoreactive fibers and terminals that is encountered in hypothalamic areas such as the medial preoptic area (MPOA), is well known to be involved in the control of pre-ovulatory LH surge. These authors studied the effects of the treatment with estrogen on the immunoreactivity of NEI and MCH in OVX animals, using the non-human primate (M. Fasscicularis). A slight increase of MCH-IR after 30 days post treatment with estrogen, along with a concurrent significant rise in NEI-IR was observed. A three-fold increase in MCH and NEI –Ir was seen at 72 h post estrogen treatment, compared with the amount of both peptides-ir at 48h post treatment. These facts suggest the possible involvement of these peptides in the regulation of the pre-ovulatory mid- cycle LH surge in primates [19].
The MCH receptors are classified into five subtypes: MC1-R to MC5-R [28]. MCH stimulates gonadotropin-releasing hormone (GnRH) release from hypothalamic explants, and it is interesting to note that MCH affects the release of LH [23] in the female rat. When MCH was injected bilaterally into the rostral preoptic area (rPOA) or medial preoptic area (mPOA) of estrogen-primed ovariectomized rats, LH release was stimulated. Two MCH receptors are involved in the MCH effect. The stimulatory action of MCH in the rPOA was inhibited by administration of antagonists for either MC-1 R or MC-5R, indicating that both ones, MC-1R and MC-5 are involved in the central control of GnRH release by MCH [19-27].
3. The effect of neuropeptide EI on LH regulation
For this study we used male and female rats, aged 10-14 weeks which were bred in our laboratory and maintained with food and water ad libitum, with a cycle of 14h/ light/10h dark and a temperature controlled environment (22±2ºC). The animal procedures were consistent with the standards established by the National Institutes of Health Guide for the Care and Use of Laboratory Animals (1996) and the AmericanVeterinarian Guidelines of Eutanasia.
The first evidence, on the effects of NEI on LH regulation was provided by Attademo et al [1], using male and ovariectomized rats treated with estrogen benzoate (10 ug) and low doses of progesterone (40ug). These animals revealed the following: when male rats were treated with intraventricular injections of NEI (1ug/1ul), the peptide induced an increase of serum LH concentration throughout the entire period studied (10-90 min). At 90 min the serum LH slightly decreased, possibly signaling initiation of the recovery of normal LH serum levels. Control rats injected with artificial cerebrospinal fluid showed practically no changes on serum LH concentrations (Fig. 3). It was also possible to see the NEI effect on ovariectomized female rats treated with estrogen plus progesterone, by using a low dose of progesterone to permit the visualization of modifications in the LH surge. Again, the neuropeptide increased LH release compared with control animals (Fig.4)\n\t\t\t
The fact that the effect of NEI may be mediated by the noradrenergic system must be taken into consideration. The peptide is known to modify DA and NA in the nucleus accumbens and caudate putamen during grooming behavior and locomotion activity [4]. As NEI behaves similarly to α-MSH, it is important to note that there is some relation between MCH-NEI and α-MSH, indicating that all three peptides are associated in a complex inter-relationship [1].
Figure 3.
Effect of icv administration of NEI or ACSF (controls) on plasma LH concentration in male rats. Blood samples were collected at 0 (before NEI or ACSF injection), 10, 30, 60 and 90 min post-injection. Bars represent the mean ± S.E.M. ∗P < 0.05 compared to controls. - Reproduced from Attademo et al, 2004, with permission from Peptides.
Figure 4.
Time course of LH release in CHR-OVX-EB-progesterone treated female rats in the presence (◦) or absence (•) of NEI. CHR-OVX rats were subcutaneously injected with 10 ug EB and 3 days later with 40 ug progesterone at 13:00 h. On the same day of the progesterone injection, the animals were injected icv 1 ug/ul of NEI or ASCF (controls) at 12:00 and 14:00 h. Blood samples were obtained between 15:00 and 21:00 h via the jugular vein, and the plasma levels of LH were measured. The plotted values represent the mean ± S.E.M. (n = 6). ∗P < 0.05 compared to controls. - Reproduced from Attademo et al, 2004, with permission from Peptides.
4. Distribution of NEI immunoreactivity
In this study, we described the anatomical substrate underlying the NEI effect of inducing LH secretion, using techniques of double and triple label immunohitochemistry, as well as dual label immunofluorescence. A group of female rats were perfused on day 15 postovariectomy; a second group received10 µg of estradiol benzoate and were perfused two days later and a third group received 10 µg of estradiol benzoate and two days later 40 µg of progesterone and were perfused 5 h after treatment. To mimic the manipulation of the animals, we used a fourth group of ovariectomized rats treated with sesame oil, and also used female intact rats at proestrus and diestrus [28].
Using these techniques, we were able to obtain the following results:
NEI-ir neurons were observed in the medial ZI, in the perifornix at the tuberal hypothalamic level and in the lateral hypothalamus. Fibers were distributed throughout the forebrain, including areas related to reproductive control and LH secretion. We observed a dense number of NEI-ir fibers in the medial septal nucleus, the diagonal band of Broca, the environs of OVLT, the preoptic area and in the internal layer of the median eminence (Fig. 5).
All fibers seen in these areas show varicosities and terminal-like structures. NEI and terminal-like structures were in close apposition with portal blood vessels and GnRH neurons expressing Fos (Fig. 6 A and B)
Figure 5.
Distribution of NEI-ir fibers in forebrain areas related to reproduction. Bright-field photomicrographs of reference sections with thionine staining showing the OVLT ( A ) and AVPV ( C ). Dark-field photomicrographs showing the distribution of NEI-ir fibers in the environs of the OVLT ( B ) and in the AVPV ( D ). E Bright-field photomicrograph showing the distribution of NEI-ir fibers in the median eminence (ME). F Dark-field photomicrograph of the same section showing the distribution of NEIir fibers in the ME. MEi = Internal layer of the median eminence; MEe = external layer of the median eminence; ox = optic chiasm; 3v = third ventricle; ac = anterior commissure; DB = nucleus of the diagonal band; MPO = medial preoptic nucleus; MS = medial septal nucleus. Scale bar: 400 μm ( A–D ), 200 μm ( E, F ). - Reproduced from Attademo et al, 2006, with permission from Neuroendocrinology.
Figure 6.
Alternative pathways for NEI induction of LH secretion. A, B Bright-field photomicrographs showing NEI-ir varicosities and terminal-like structures in the median eminence (ME). Note the close proximity to blood vessels (arrows). C Bright-field photomicrograph showing NEI-ir varicosities in close apposition with AVPV neurons expressing Fos (Fos-ir) in the afternoon of the proestrus day (arrows). D Bright-field photomicrograph showing NEI-ir fibers in close apposition with GnRH-ir neurons expressing Fos in the afternoon of the proestrus day (arrows). E Bright-field photomicrograph showing NEI-ir varicosities (in black) in close apposition with GnRH-ir fibers (in light brown) in the ME. F Fluorescence photomicrograph showing the close association between NEI-ir (in green, AlexaFluor 488) and TH-immunoreactive (TH-ir, in red, AlexaFluor 594) neurons in the medial zona incerta (ZIm). G Fluorescence photomicrograph showing TH-ir fibers in close apposition with NEI-ir neurons in the ZIm (arrow). v = Blood vessel; MPO = medial preoptic nucleus. Scale bar: 50 μm ( A–E, G ); 100 μm ( F ). - Reproduced from Attademo et al, 2006, with permission from Neuroendocrinology.
5. NEI-ir fibers innervate GnRH and AVPV neurons expresing fos
We observed an increased expression of Fos immunoreactivity in the anteroventral periventricular nucleus (AVPV) neurons of rats perfused during the afternoon of the day of proestrus, as well as in ovariectomized rats treated with estradiol benzoate plus progesterone, as described by others [30, 31]. Close to a 10% of the AVPV neurons expressing Fos receive NEI fibers in close apposition. Very little Fos immunoreactivity was observed in rats perfused in the afternoon of the diestrus day, for either of the following conditions: ovariectomized treated with estradiol benzoate or ovariectomized treated with sesame oil. In addition, all groups, showed a sparse distribution of Fos immunoreactivity in the medial zone incerta and in the LHA, whereas co-localization of NEI fibers with any of these cells was not observed (Fig. 6 D).
It has been reported that Fos protein is expressed in a portion of GnRH neurons that are active in the afternoon of the proestrus day, as well as in the GnRH neurons of ovariectomized rats treated with estradiol benzoate plus progesterone [31-33]. Based on these results, we investigated the pattern of NEI innervations in areas that expressed GnRH neurons in these animals. Most of the neurons expressing Fos were found to be in the vicinity of the organum vasculosum of the lamina terminalis (OVLT) and of the preoptic area. NEI fibers were also found in the median eminence, and NEI and GnRH varicosities presented similar distribution, thus revealing close apposition between them. Immediately before and during LH surge (on the afternoon of the proestrus day or following treatment with estrogen plus progesterone), only a portion of the GnRH neurons expressed Fos [30,31,33,34-36].
AVPV, is a nucleus that also expresses Fos in the afternoon of the proestrus day and has been widely implicated in the control of reproduction [32], with Fos expression indicating neuronal response [36]. However, in rats perfused on the diestrus day or in ovariectomized rats receiving estradiol benzoate or sesame oil, we did not find Fos expression in GnRH neurons. The pattern of distribution of NEI-ir and GnRH ir in the median eminence, showed NEI fibers to be denser in the internal layer than in the external one. Nevertheless, in the external layer, NEI and GnRH varicosities presented similar distributions, revealing an apparent close apposition between them. In the present study, we labeled distinctive cell compartments by using dual and triple-label immunohistochemistry which revealed NEI-ir fibers to be innervating the Fos-positive neurons in the AVPV, as well as the GnRH neurons positive for Fos immunoreactivity. In these experiments we labeled distinct cell compartments (cytoplasm, nuclei and terminals) using various antisera, all raised in rabbit. In the control tests, the second or third antisera were omitted, and no reaction was evident. This indicates that the observed labeling of cell bodies, fibers or both was not the result of cross-reactivity of the secondary antibody. In addition, evaluation of the data under light microscopy at a high magnification revealed only a suggestion of synaptic contact (Fig 6 A-B).
6. NEI innervation of GnRH neurons
Our results indicated that NEI fibers were in close apposition with GnRH neurons expressing Fos in the afternoon of the proestrus day. Since Fos protein expression in GnRH neurons increases in parallel with rises in the plasma LH levels [27, 31, 32], it can be assumed that these neurons project to the median eminence and induce LH secretion during proestrus. Therefore, we can suggest that NEI (through projections to a subset of GnRH neurons) modulated GnRH activity and, consequently, LH surges.
The role that NEI plays in GnRH secretion has not been investigated. However, NEI varicosities in some parts of the median eminence display a pattern of distribution similar to that of GnRH, revealing a possible effect on the modulation of GnRH secretion directly at the terminals. This may represent one of the mechanisms by which intracerebroventricular administration of NEI causes an increase in LH secretion (Fig. 6 E)
Experiments using in vitro preparations or intracerebroventricular injections have shown that various neurotransmitters can regulate gonadotropin release [38-40]. One such neurotransmitters is the cocaine- and amphetamine- regulated transcript (CART) peptide, which has been shown to increase the GnRH pulse amplitude in cycling female rats and to decrease GnRH pulse intervals in prepubertal rats [41-43]. These effects can be achieved through direct CART innervation of the GnRH neurons [39, 40]. Interestingly, in the medial ZI and lateral hypothalamus, MCH/NEI neurons coexpress CART [44, 46]. In the present study, we did not explore the origins of NEI innervation of GnRH neurons or areas related to reproductive behavior. However, it is intriguing that the number of NEI-ir neurons in ovariectomized rats was increased only in the medial ZI, a brain region that projects to the AVPV and GnRH- containing areas [47,48], as well as to the circumventricular organs, probably including the median eminence [44, 45]. This result is in agreement with those of other studies in which ppMCH mRNA expression was found to be greater in the medial ZI of untreated ovariectomized rats than in that of ovariectomized rats primed with estradiol benzoate or estradiol benzoate plus progesterone.
7. In vitro studies
Taken into account the above results, we decided to investigate whether NEI could act directly at the pituitary level by modulating hormone secretion. With this purpose the effects of NEI were studied in pituitary cell cultures from female rats on the release of several pituitary hormones (GH, LH and prolactin). Furthermore, the ability of NEI to activate pituitary cells was evaluated by electron microscopy and immunocitochemistry [49], and finally, the ability of NEI to potentiate GnRH-induced LH release was tested. The study of the effects of physiological stimuli involved in the regulation of pituitary hormone secretion was facilitated by the availability of a system consisting of a suspension of single dispersed pituitary cells, in which the cell functions were essentially the same as in situ [50]. For this study we used female rats in order to obtain a primary culture. The results were as follows:
NEI induced a fast release of LH in the culture cell media. In addition, there were differences in the LH levels obtained for the various time periods of hormonal stimuli assayed, which were closely associated with the doses applied. The lowest dose of NEI (100x 10-8M) induced a significant increase of LH secretion after 2h of stimulus, achieving maximum response after 4h of NEI treatment. At this time, the LH levels almost reached a five-fold increase over controls and then maintained these values (Fig.7). In spite of NEI being effective in stimulating LH secretion, none of the assayed doses were capable of significantly promoting FSH secretion from gonadotrophs. In addition, no significant increases were observed on prolactin or GH secretion, at doses ranging from 100x10-8 to 400x10-8 M NEI in primary cell cultures, thus confirming the specificity of NEI stimuli on LH secretion (Table 1). To determine whether or not NEI was able to synergize with GnRH in stimulating LH release, pituitary cells were simultaneously incubated with GnRH (0.1 or 1x10-9M) and different concentrations of NEI (1, 10 or 100x10-8M) for 3 h and then the media was collected and tested for LH by RIA (Fig. 8). Although NEI at the dose of 10x10-8M had no effect on LH secretion, GnRH 1x10-9M plus NEI 10x10-8M induced a slight but significant increase in LH concentrations (16%; p< 0.01). A combined treatment with the highest doses of both NEI and GnRH significantly stimulated the secretory response, which was more effective than that observed with the same dose of GnRH alone [49] (Fig.8).
When pituitary cells from female rats were cultured, different types of secretory cells were observed (Fig. 9). These were identifiable by their ultra structural characteristics, essentially by the profile of the secretory granules, which constituted a distinctive feature. The most frequent populations observed were lactotroph and somatotroph cells, which were in close contact with gonadotrph cells. In the control group, the lactotroph and somatotroph cells have numerous polymorphic and round mature secretory granules respectively. These also had high electron densities and were stored in the cytoplasm. The gonadotroph were
Figure 7.
Time-course study of the effects of NEI on LH secretion in the culture media. The cell cultures were treated with NEI 100 or 400 x 10-8 M for 1–5 h, in serum free conditions. The data are represented as mean ± S.E.M. of three independent experiments. Data were evaluated by the ANOVA–Fisher test; *p < 0.01 vs Control group. – Reproduced from De Paul et al, 2009, with permission from Peptides.
Table 1.
Time-course study of the effects of NEI on FSH, PRL and GH secretion accumulated in the culture media (ng/ml of culture medium). Pituitary cells were treated with NEI 100 or 400×10-8M for 1–5 h. The data are shown as the mean ±S.E.M. of three independent experiments and were evaluated by the ANOVA–Fisher test.
Figure 8.
NEI and GnRH combined treatments on LH secretion. The presence of 10 and 100 x 10-8 M NEI in the culture media for 3 h promoted a significant increase in LH release stimulated by GnRH 1 x10-9 M. Data are shown as the mean ± S.E.M. of three independent experiments. ANOVA–Fisher test; *p < 0.01 vs control group; **p < 0.01 vs GnRH 1 x 10-9 M. – Reproduced from De Paul et al, 2009, with permission from Peptides.
characterized by a conspicuous accumulation of round secretory granules of two different sizes and by electron densities in the cytoplasm. The most abundant granules were about 150nm in diameter and filled with homogenous material, whereas the others less frequent but larger in size (about 400nm) (Fig. 10 A and B).Stimulation with NEI (100 x 10-8 and 400 x 10 -8) for 2 and 4 hour, promoted several subtle structural changes, particularly in the
Figure 9.
A) Electron micrograph of a pituitary cell culture from control rats illustrating different pituitary cell populations. Two lactotroph cells (L) exhibit an accumulation of large and polymorphic mature secretory granules (about 500– 900 nm in diameter) in the cytoplasm. The somatotroph cell (S) can be easily recognized by the mature, round GH secretory granules ranging from 200 to 350 nm in diameter and scattered throughout the cytoplasm. In close contact with both secretory cell types, a gonadotroph cell (G) displays small (ssg) and large (lsg) round secretory granules (about 150 or 400 nm in diameter respectively, with different electron densities, homogeneously disseminated in the cytoplasm (m = mitochondria, N = nucleus). Bar: 1 μm. (B) Gonadotroph cell specifically immunostained for LH. The cytoplasm shows a noticeable accumulation of characteristic small (ssg) and large (lsg) round secretory granules (m = mitochondria, N = nucleus). Bar: 0.5 μm. – Reproduced from De Paul et al, 2009, with permission from Peptides.
Figure 10.
A) Electron microscopy of two cultured gonadotroph cells treated with 400 x 10-8 M NEI for 4 h. The cytoplasm contains a remarkably well developed rough endoplasmic reticulum (RER) and Golgi complex (GC) and also scarce small secretory granules (ssg) that are mostly in contact with the plasma membrane. Bar: 0.5 μm. (B) Electron micrograph of a cultured gonadotroph cell (G) after exposition to 100 x 10-8 M NEI for 2 h which is shown exhibiting small secretory granules (ssg) mobilized toward the plasmalemma, where they will then be subsequently discharged by exocytosis. Bar: 0.5 mm. Inset: small round secretory granules from a gonadotroph cell specifically identified by immunocytochemistry for LH. Bar: 0.5 mm. (C) Detail of two adjacent gonadotroph cells after NEI treatment displaying evidence of secretory activity. The secretory granules are aligned alongside the cell membrane and are in the process of exocytosis (arrows). Bar: 0.5 μm. – Reproduced from De Paul et al, 2009, with permission from Peptides.
gonadotroph cell population. For this cell type, the most prominent changes consisted of a striking development of the rough endoplasmic reticulum (RER) and Golgi complex (Fig 10B) when compared to the control group. Many secretory granules were located to the cell membrane and presented images of exocytosis after NEI treatment (Fig. 10 C) Other pituitary cell populations, the lactotrophs, thyrotrophs and somatotrophs, did not exhibit any features indicating a significant activation of hormone release after NEI treatment.
The present results were the first demonstration of a specific and direct action of NEI in cultured pituitary cells without modifying other pituitary hormones. Moreover, the analysis of the electron microscope images taken 2 and 4h after NEI treatment was indicative of the stimulation of LH release occurring at these times.
From the present study, it is possible to conclude that NEI is effective when injected into the brain to release LH in male and female rats. The anatomical substrate underlying this effect was identified using combined methods of immunohistochemistry. A schematic representation of the proposed pathways by which NEI participates in LH secretion is depicted in Figure 11.
NEI is also capable of inducing a marked release of LH without modifying the other pituitary hormones in the pituitary cultured cells. There is an interaction between NEI and GnRH in vivo and in vitro.
Figure 11.
Schematic representation of the proposed pathways by which NEI participates in LH secretion. NEI-ir neurons in the medial zona incerta (ZIm) or LHA receive dopaminergic innervation from TH neurons located in the ZIm and project directly to the median eminence or to GnRH neurons in the preoptic area. In addition, NEI might modulate LH secretion by innervating the AVPV. – Reproduced from Attademo et al, 2006, with permission from Neuroendocrinology.
Acknowledgement
First of all I wish to thank to Dr. Paul Hobson for critical reading of this manuscript. I also wish to thank Dr. Jorge Paván, Miss Laura Alazraki and Miss Marina Juarez for editing the manuscript. Thanks for the grants from CONICET; SECyT and FONCyT. MEC is member of CONICET.
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The effect of neuropeptide EI on LH regulation",level:"1"},{id:"sec_4",title:"4. Distribution of NEI immunoreactivity ",level:"1"},{id:"sec_5",title:"5. NEI-ir fibers innervate GnRH and AVPV neurons expresing fos",level:"1"},{id:"sec_6",title:"6. NEI innervation of GnRH neurons",level:"1"},{id:"sec_7",title:"7. In vitro studies",level:"1"},{id:"sec_7_2",title:"Acknowledgement",level:"2"}],chapterReferences:[{id:"B1",body:'AttademoA. MSanchez-borzoneMLasagaMCelisM. EIntracerebroventicular injection or neuropeptide El increases serum LH in male and female rats. Peptides 20042519959\n\t\t\t'},{id:"B2",body:'BerberianVSanchezM. SCelisM. EParticipation of the cholinergic system in the exesesive grooming behavior induced by neuropeptide (N) glutamic acid (E) isoleucine (I) amide (NEI). Neurochem Res 20022717131717'},{id:"B3",body:'SanchezMBakerB. 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Martín-Acebes, Ángela Vázquez-Calvo, Flavia Caridi, Juan-Carlos Saiz and Francisco Sobrino",authors:[{id:"141163",title:"Dr.",name:"Francisco",middleName:null,surname:"Sobrino",fullName:"Francisco Sobrino",slug:"francisco-sobrino"}]},{id:"42116",title:"The Role of Altered Lipid Metabolism in Septic Myocardial Dysfunction",slug:"the-role-of-altered-lipid-metabolism-in-septic-myocardial-dysfunction",signatures:"Luca Siracusano and Viviana Girasole",authors:[{id:"139192",title:"Prof.",name:"Luca",middleName:null,surname:"Siracusano",fullName:"Luca Siracusano",slug:"luca-siracusano"},{id:"139193",title:"Dr.",name:"Viviana",middleName:null,surname:"Girasole",fullName:"Viviana Girasole",slug:"viviana-girasole"}]},{id:"42108",title:"Lipids as Markers of Induced Resistance in Wheat: A Biochemical and Molecular Approach",slug:"lipids-as-markers-of-induced-resistance-in-wheat-a-biochemical-and-molecular-approach",signatures:"Christine Tayeh, Béatrice Randoux, Frédéric Laruelle, Natacha Bourdon, Delphine Renard-Merlier and Philippe Reignault",authors:[{id:"140626",title:"Prof.",name:"Philippe",middleName:null,surname:"Reignault",fullName:"Philippe Reignault",slug:"philippe-reignault"}]},{id:"42122",title:"Jasmonate Biosynthesis, Perception and Function in Plant Development and Stress Responses",slug:"jasmonate-biosynthesis-perception-and-function-in-plant-development-and-stress-responses",signatures:"Yuanxin Yan, Eli Borrego and Michael V. Kolomiets",authors:[{id:"141200",title:"Prof.",name:"Michael",middleName:null,surname:"Kolomiets",fullName:"Michael Kolomiets",slug:"michael-kolomiets"},{id:"141211",title:"Dr.",name:"Yuanxin",middleName:null,surname:"Yan",fullName:"Yuanxin Yan",slug:"yuanxin-yan"}]},{id:"42119",title:"The Effect of Probiotics on Lipid Metabolism",slug:"the-effect-of-probiotics-on-lipid-metabolism",signatures:"Yong Zhang and Heping Zhang",authors:[{id:"141580",title:"Mr.",name:"Yong",middleName:null,surname:"Zhang",fullName:"Yong Zhang",slug:"yong-zhang"}]}]}]},onlineFirst:{chapter:{type:"chapter",id:"68505",title:"Research Design and Methodology",doi:"10.5772/intechopen.85731",slug:"research-design-and-methodology",body:'
1. Introduction
Research methodology is the path through which researchers need to conduct their research. It shows the path through which these researchers formulate their problem and objective and present their result from the data obtained during the study period. This research design and methodology chapter also shows how the research outcome at the end will be obtained in line with meeting the objective of the study. This chapter hence discusses the research methods that were used during the research process. It includes the research methodology of the study from the research strategy to the result dissemination. For emphasis, in this chapter, the author outlines the research strategy, research design, research methodology, the study area, data sources such as primary data sources and secondary data, population consideration and sample size determination such as questionnaires sample size determination and workplace site exposure measurement sample determination, data collection methods like primary data collection methods including workplace site observation data collection and data collection through desk review, data collection through questionnaires, data obtained from experts opinion, workplace site exposure measurement, data collection tools pretest, secondary data collection methods, methods of data analysis used such as quantitative data analysis and qualitative data analysis, data analysis software, the reliability and validity analysis of the quantitative data, reliability of data, reliability analysis, validity, data quality management, inclusion criteria, ethical consideration and dissemination of result and its utilization approaches. In order to satisfy the objectives of the study, a qualitative and quantitative research method is apprehended in general. The study used these mixed strategies because the data were obtained from all aspects of the data source during the study time. Therefore, the purpose of this methodology is to satisfy the research plan and target devised by the researcher.
2. Research design
The research design is intended to provide an appropriate framework for a study. A very significant decision in research design process is the choice to be made regarding research approach since it determines how relevant information for a study will be obtained; however, the research design process involves many interrelated decisions [1].
This study employed a mixed type of methods. The first part of the study consisted of a series of well-structured questionnaires (for management, employee’s representatives, and technician of industries) and semi-structured interviews with key stakeholders (government bodies, ministries, and industries) in participating organizations. The other design used is an interview of employees to know how they feel about safety and health of their workplace, and field observation at the selected industrial sites was undertaken.
Hence, this study employs a descriptive research design to agree on the effects of occupational safety and health management system on employee health, safety, and property damage for selected manufacturing industries. Saunders et al. [2] and Miller [3] say that descriptive research portrays an accurate profile of persons, events, or situations. This design offers to the researchers a profile of described relevant aspects of the phenomena of interest from an individual, organizational, and industry-oriented perspective. Therefore, this research design enabled the researchers to gather data from a wide range of respondents on the impact of safety and health on manufacturing industries in Ethiopia. And this helped in analyzing the response obtained on how it affects the manufacturing industries’ workplace safety and health. The research overall design and flow process are depicted in Figure 1.
Figure 1.
Research methods and processes (author design).
3. Research methodology
To address the key research objectives, this research used both qualitative and quantitative methods and combination of primary and secondary sources. The qualitative data supports the quantitative data analysis and results. The result obtained is triangulated since the researcher utilized the qualitative and quantitative data types in the data analysis. The study area, data sources, and sampling techniques were discussed under this section.
3.1 The study area
According to Fraenkel and Warren [4] studies, population refers to the complete set of individuals (subjects or events) having common characteristics in which the researcher is interested. The population of the study was determined based on random sampling system. This data collection was conducted from March 07, 2015 to December 10, 2016, from selected manufacturing industries found in Addis Ababa city and around. The manufacturing companies were selected based on their employee number, established year, and the potential accidents prevailing and the manufacturing industry type even though all criterions were difficult to satisfy.
3.2 Data sources
3.2.1 Primary data sources
It was obtained from the original source of information. The primary data were more reliable and have more confidence level of decision-making with the trusted analysis having direct intact with occurrence of the events. The primary data sources are industries’ working environment (through observation, pictures, and photograph) and industry employees (management and bottom workers) (interview, questionnaires and discussions).
3.2.2 Secondary data
Desk review has been conducted to collect data from various secondary sources. This includes reports and project documents at each manufacturing sectors (more on medium and large level). Secondary data sources have been obtained from literatures regarding OSH, and the remaining data were from the companies’ manuals, reports, and some management documents which were included under the desk review. Reputable journals, books, different articles, periodicals, proceedings, magazines, newsletters, newspapers, websites, and other sources were considered on the manufacturing industrial sectors. The data also obtained from the existing working documents, manuals, procedures, reports, statistical data, policies, regulations, and standards were taken into account for the review.
In general, for this research study, the desk review has been completed to this end, and it had been polished and modified upon manuals and documents obtained from the selected companies.
4. Population and sample size
4.1 Population
The study population consisted of manufacturing industries’ employees in Addis Ababa city and around as there are more representative manufacturing industrial clusters found. To select representative manufacturing industrial sector population, the types of the industries expected were more potential to accidents based on random and purposive sampling considered. The population of data was from textile, leather, metal, chemicals, and food manufacturing industries. A total of 189 sample sizes of industries responded to the questionnaire survey from the priority areas of the government. Random sample sizes and disproportionate methods were used, and 80 from wood, metal, and iron works; 30 from food, beverage, and tobacco products; 50 from leather, textile, and garments; 20 from chemical and chemical products; and 9 from other remaining 9 clusters of manufacturing industries responded.
4.2 Questionnaire sample size determination
A simple random sampling and purposive sampling methods were used to select the representative manufacturing industries and respondents for the study. The simple random sampling ensures that each member of the population has an equal chance for the selection or the chance of getting a response which can be more than equal to the chance depending on the data analysis justification. Sample size determination procedure was used to get optimum and reasonable information. In this study, both probability (simple random sampling) and nonprobability (convenience, quota, purposive, and judgmental) sampling methods were used as the nature of the industries are varied. This is because of the characteristics of data sources which permitted the researchers to follow the multi-methods. This helps the analysis to triangulate the data obtained and increase the reliability of the research outcome and its decision. The companies’ establishment time and its engagement in operation, the number of employees and the proportion it has, the owner types (government and private), type of manufacturing industry/production, types of resource used at work, and the location it is found in the city and around were some of the criteria for the selections.
The determination of the sample size was adopted from Daniel [5] and Cochran [6] formula. The formula used was for unknown population size Eq. (1) and is given as
E1
where n = sample size, Z = statistic for a level of confidence, P = expected prevalence or proportion (in proportion of one; if 50%, P = 0.5), and d = precision (in proportion of one; if 6%, d = 0.06). Z statistic (Z): for the level of confidence of 95%, which is conventional, Z value is 1.96. In this study, investigators present their results with 95% confidence intervals (CI).
The expected sample number was 267 at the marginal error of 6% for 95% confidence interval of manufacturing industries. However, the collected data indicated that only 189 populations were used for the analysis after rejecting some data having more missing values in the responses from the industries. Hence, the actual data collection resulted in 71% response rate. The 267 population were assumed to be satisfactory and representative for the data analysis.
4.3 Workplace site exposure measurement sample determination
The sample size for the experimental exposure measurements of physical work environment has been considered based on the physical data prepared for questionnaires and respondents. The response of positive were considered for exposure measurement factors to be considered for the physical environment health and disease causing such as noise intensity, light intensity, pressure/stress, vibration, temperature/coldness, or hotness and dust particles on 20 workplace sites. The selection method was using random sampling in line with purposive method. The measurement of the exposure factors was done in collaboration with Addis Ababa city Administration and Oromia Bureau of Labour and Social Affair (AACBOLSA). Some measuring instruments were obtained from the Addis Ababa city and Oromia Bureau of Labour and Social Affair.
5. Data collection methods
Data collection methods were focused on the followings basic techniques. These included secondary and primary data collections focusing on both qualitative and quantitative data as defined in the previous section. The data collection mechanisms are devised and prepared with their proper procedures.
5.1 Primary data collection methods
Primary data sources are qualitative and quantitative. The qualitative sources are field observation, interview, and informal discussions, while that of quantitative data sources are survey questionnaires and interview questions. The next sections elaborate how the data were obtained from the primary sources.
5.1.1 Workplace site observation data collection
Observation is an important aspect of science. Observation is tightly connected to data collection, and there are different sources for this: documentation, archival records, interviews, direct observations, and participant observations. Observational research findings are considered strong in validity because the researcher is able to collect a depth of information about a particular behavior. In this dissertation, the researchers used observation method as one tool for collecting information and data before questionnaire design and after the start of research too. The researcher made more than 20 specific observations of manufacturing industries in the study areas. During the observations, it found a deeper understanding of the working environment and the different sections in the production system and OSH practices.
5.1.2 Data collection through interview
Interview is a loosely structured qualitative in-depth interview with people who are considered to be particularly knowledgeable about the topic of interest. The semi-structured interview is usually conducted in a face-to-face setting which permits the researcher to seek new insights, ask questions, and assess phenomena in different perspectives. It let the researcher to know the in-depth of the present working environment influential factors and consequences. It has provided opportunities for refining data collection efforts and examining specialized systems or processes. It was used when the researcher faces written records or published document limitation or wanted to triangulate the data obtained from other primary and secondary data sources.
This dissertation is also conducted with a qualitative approach and conducting interviews. The advantage of using interviews as a method is that it allows respondents to raise issues that the interviewer may not have expected. All interviews with employees, management, and technicians were conducted by the corresponding researcher, on a face-to-face basis at workplace. All interviews were recorded and transcribed.
5.1.3 Data collection through questionnaires
The main tool for gaining primary information in practical research is questionnaires, due to the fact that the researcher can decide on the sample and the types of questions to be asked [2].
In this dissertation, each respondent is requested to reply to an identical list of questions mixed so that biasness was prevented. Initially the questionnaire design was coded and mixed up from specific topic based on uniform structures. Consequently, the questionnaire produced valuable data which was required to achieve the dissertation objectives.
The questionnaires developed were based on a five-item Likert scale. Responses were given to each statement using a five-point Likert-type scale, for which 1 = “strongly disagree” to 5 = “strongly agree.” The responses were summed up to produce a score for the measures.
5.1.4 Data obtained from experts’ opinion
The data was also obtained from the expert’s opinion related to the comparison of the knowledge, management, collaboration, and technology utilization including their sub-factors. The data obtained in this way was used for prioritization and decision-making of OSH, improving factor priority. The prioritization of the factors was using Saaty scales (1–9) and then converting to Fuzzy set values obtained from previous researches using triangular fuzzy set [7].
5.1.5 Workplace site exposure measurement
The researcher has measured the workplace environment for dust, vibration, heat, pressure, light, and noise to know how much is the level of each variable. The primary data sources planned and an actual coverage has been compared as shown in Table 1.
Table 1.
Planned versus actual coverage of the survey.
The response rate for the proposed data source was good, and the pilot test also proved the reliability of questionnaires. Interview/discussion resulted in 87% of responses among the respondents; the survey questionnaire response rate obtained was 71%, and the field observation response rate was 90% for the whole data analysis process. Hence, the data organization quality level has not been compromised.
This response rate is considered to be representative of studies of organizations. As the study agrees on the response rate to be 30%, it is considered acceptable [8]. Saunders et al. [2] argued that the questionnaire with a scale response of 20% response rate is acceptable. Low response rate should not discourage the researchers, because a great deal of published research work also achieves low response rate. Hence, the response rate of this study is acceptable and very good for the purpose of meeting the study objectives.
5.1.6 Data collection tool pretest
The pretest for questionnaires, interviews, and tools were conducted to validate that the tool content is valid or not in the sense of the respondents’ understanding. Hence, content validity (in which the questions are answered to the target without excluding important points), internal validity (in which the questions raised answer the outcomes of researchers’ target), and external validity (in which the result can generalize to all the population from the survey sample population) were reflected. It has been proved with this pilot test prior to the start of the basic data collections. Following feedback process, a few minor changes were made to the originally designed data collect tools. The pilot test made for the questionnaire test was on 10 sample sizes selected randomly from the target sectors and experts.
5.2 Secondary data collection methods
The secondary data refers to data that was collected by someone other than the user. This data source gives insights of the research area of the current state-of-the-art method. It also makes some sort of research gap that needs to be filled by the researcher. This secondary data sources could be internal and external data sources of information that may cover a wide range of areas.
Literature/desk review and industry documents and reports: To achieve the dissertation’s objectives, the researcher has conducted excessive document review and reports of the companies in both online and offline modes. From a methodological point of view, literature reviews can be comprehended as content analysis, where quantitative and qualitative aspects are mixed to assess structural (descriptive) as well as content criteria.
A literature search was conducted using the database sources like MEDLINE; Emerald; Taylor and Francis publications; EMBASE (medical literature); PsycINFO (psychological literature); Sociological Abstracts (sociological literature); accident prevention journals; US Statistics of Labor, European Safety and Health database; ABI Inform; Business Source Premier (business/management literature); EconLit (economic literature); Social Service Abstracts (social work and social service literature); and other related materials. The search strategy was focused on articles or reports that measure one or more of the dimensions within the research OSH model framework. This search strategy was based on a framework and measurement filter strategy developed by the Consensus-Based Standards for the Selection of Health Measurement Instruments (COSMIN) group. Based on screening, unrelated articles to the research model and objectives were excluded. Prior to screening, researcher (principal investigator) reviewed a sample of more than 2000 articles, websites, reports, and guidelines to determine whether they should be included for further review or reject. Discrepancies were thoroughly identified and resolved before the review of the main group of more than 300 articles commenced. After excluding the articles based on the title, keywords, and abstract, the remaining articles were reviewed in detail, and the information was extracted on the instrument that was used to assess the dimension of research interest. A complete list of items was then collated within each research targets or objectives and reviewed to identify any missing elements.
6. Methods of data analysis
Data analysis method follows the procedures listed under the following sections. The data analysis part answered the basic questions raised in the problem statement. The detailed analysis of the developed and developing countries’ experiences on OSH regarding manufacturing industries was analyzed, discussed, compared and contrasted, and synthesized.
6.1 Quantitative data analysis
Quantitative data were obtained from primary and secondary data discussed above in this chapter. This data analysis was based on their data type using Excel, SPSS 20.0, Office Word format, and other tools. This data analysis focuses on numerical/quantitative data analysis.
Before analysis, data coding of responses and analysis were made. In order to analyze the data obtained easily, the data were coded to SPSS 20.0 software as the data obtained from questionnaires. This task involved identifying, classifying, and assigning a numeric or character symbol to data, which was done in only one way pre-coded [9, 10]. In this study, all of the responses were pre-coded. They were taken from the list of responses, a number of corresponding to a particular selection was given. This process was applied to every earlier question that needed this treatment. Upon completion, the data were then entered to a statistical analysis software package, SPSS version 20.0 on Windows 10 for the next steps.
Under the data analysis, exploration of data has been made with descriptive statistics and graphical analysis. The analysis included exploring the relationship between variables and comparing groups how they affect each other. This has been done using cross tabulation/chi square, correlation, and factor analysis and using nonparametric statistic.
6.2 Qualitative data analysis
Qualitative data analysis used for triangulation of the quantitative data analysis. The interview, observation, and report records were used to support the findings. The analysis has been incorporated with the quantitative discussion results in the data analysis parts.
6.3 Data analysis software
The data were entered using SPSS 20.0 on Windows 10 and analyzed. The analysis supported with SPSS software much contributed to the finding. It had contributed to the data validation and correctness of the SPSS results. The software analyzed and compared the results of different variables used in the research questionnaires. Excel is also used to draw the pictures and calculate some analytical solutions.
7. The reliability and validity analysis of the quantitative data
7.1 Reliability of data
The reliability of measurements specifies the amount to which it is without bias (error free) and hence ensures consistent measurement across time and across the various items in the instrument [8]. In reliability analysis, it has been checked for the stability and consistency of the data. In the case of reliability analysis, the researcher checked the accuracy and precision of the procedure of measurement. Reliability has numerous definitions and approaches, but in several environments, the concept comes to be consistent [8]. The measurement fulfills the requirements of reliability when it produces consistent results during data analysis procedure. The reliability is determined through Cranach’s alpha as shown in Table 2.
Table 2.
Internal consistency and reliability test of questionnaires items.
K stands for knowledge; M, management; T, technology; C, collaboration; P, policy, standards, and regulation; H, hazards and accident conditions; PPE, personal protective equipment.
7.2 Reliability analysis
Cronbach’s alpha is a measure of internal consistency, i.e., how closely related a set of items are as a group [11]. It is considered to be a measure of scale reliability. The reliability of internal consistency most of the time is measured based on the Cronbach’s alpha value. Reliability coefficient of 0.70 and above is considered “acceptable” in most research situations [12]. In this study, reliability analysis for internal consistency of Likert-scale measurement after deleting 13 items was found similar; the reliability coefficients were found for 76 items were 0.964 and for the individual groupings made shown in Table 2. It was also found internally consistent using the Cronbach’s alpha test. Table 2 shows the internal consistency of the seven major instruments in which their reliability falls in the acceptable range for this research.
7.3 Validity
Face validity used as defined by Babbie [13] is an indicator that makes it seem a reasonable measure of some variables, and it is the subjective judgment that the instrument measures what it intends to measure in terms of relevance [14]. Thus, the researcher ensured, in this study, when developing the instruments that uncertainties were eliminated by using appropriate words and concepts in order to enhance clarity and general suitability [14]. Furthermore, the researcher submitted the instruments to the research supervisor and the joint supervisor who are both occupational health experts, to ensure validity of the measuring instruments and determine whether the instruments could be considered valid on face value.
In this study, the researcher was guided by reviewed literature related to compliance with the occupational health and safety conditions and data collection methods before he could develop the measuring instruments. In addition, the pretest study that was conducted prior to the main study assisted the researcher to avoid uncertainties of the contents in the data collection measuring instruments. A thorough inspection of the measuring instruments by the statistician and the researcher’s supervisor and joint experts, to ensure that all concepts pertaining to the study were included, ensured that the instruments were enriched.
8. Data quality management
Insight has been given to the data collectors on how to approach companies, and many of the questionnaires were distributed through MSc students at Addis Ababa Institute of Technology (AAiT) and manufacturing industries’ experience experts. This made the data quality reliable as it has been continually discussed with them. Pretesting for questionnaire was done on 10 workers to assure the quality of the data and for improvement of data collection tools. Supervision during data collection was done to understand how the data collectors are handling the questionnaire, and each filled questionnaires was checked for its completeness, accuracy, clarity, and consistency on a daily basis either face-to-face or by phone/email. The data expected in poor quality were rejected out of the acting during the screening time. Among planned 267 questionnaires, 189 were responded back. Finally, it was analyzed by the principal investigator.
9. Inclusion criteria
The data were collected from the company representative with the knowledge of OSH. Articles written in English and Amharic were included in this study. Database information obtained in relation to articles and those who have OSH area such as interventions method, method of accident identification, impact of occupational accidents, types of occupational injuries/disease, and impact of occupational accidents, and disease on productivity and costs of company and have used at least one form of feedback mechanism. No specific time period was chosen in order to access all available published papers. The questionnaire statements which are similar in the questionnaire have been rejected from the data analysis.
10. Ethical consideration
Ethical clearance was obtained from the School of Mechanical and Industrial Engineering, Institute of Technology, Addis Ababa University. Official letters were written from the School of Mechanical and Industrial Engineering to the respective manufacturing industries. The purpose of the study was explained to the study subjects. The study subjects were told that the information they provided was kept confidential and that their identities would not be revealed in association with the information they provided. Informed consent was secured from each participant. For bad working environment assessment findings, feedback will be given to all manufacturing industries involved in the study. There is a plan to give a copy of the result to the respective study manufacturing industries’ and ministries’ offices. The respondents’ privacy and their responses were not individually analyzed and included in the report.
11. Dissemination and utilization of the result
The result of this study will be presented to the Addis Ababa University, AAiT, School of Mechanical and Industrial Engineering. It will also be communicated to the Ethiopian manufacturing industries, Ministry of Labor and Social Affair, Ministry of Industry, and Ministry of Health from where the data was collected. The result will also be availed by publication and online presentation in Google Scholars. To this end, about five articles were published and disseminated to the whole world.
12. Conclusion
The research methodology and design indicated overall process of the flow of the research for the given study. The data sources and data collection methods were used. The overall research strategies and framework are indicated in this research process from problem formulation to problem validation including all the parameters. It has laid some foundation and how research methodology is devised and framed for researchers. This means, it helps researchers to consider it as one of the samples and models for the research data collection and process from the beginning of the problem statement to the research finding. Especially, this research flow helps new researchers to the research environment and methodology in particular.
Conflict of interest
There is no “conflict of interest.”
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New York: John Wiley & Sons Inc; 2000'},{id:"B2",body:'Saunders M, Lewis P, Thornhill A. Research Methods for Business Student. 5th ed. Edinburgh Gate: Pearson Education Limited; 2009'},{id:"B3",body:'Miller P. Motivation in the Workplace. Work and Organizational Psychology. Oxford: Blackwell Publishers; 1991'},{id:"B4",body:'Fraenkel FJ, Warren NE. How to Design and Evaluate Research in Education. 4th ed. New York: McGraw-Hill; 2002'},{id:"B5",body:'Danniel WW. Biostatist: A Foundation for Analysis in the Health Science. 7th ed. New York: John Wiley & Sons; 1999'},{id:"B6",body:'Cochran WG. Sampling Techniques. 3rd ed. New York: John Wiley & Sons; 1977'},{id:"B7",body:'Saaty TL. The Analytical Hierarchy Process. Pittsburg: PWS Publications; 1990'},{id:"B8",body:'Sekaran U, Bougie R. Research Methods for Business: A Skill Building Approach. 5th ed. New Delhi: John Wiley & Sons, Ltd; 2010. pp. 1-468'},{id:"B9",body:'Luck DJ, Rubin RS. Marketing Research. 7th ed. New Jersey: Prentice-Hall International; 1987'},{id:"B10",body:'Wong TC. Marketing Research. Oxford, UK: Butterworth-Heinemann; 1999'},{id:"B11",body:'Cronbach LJ. Coefficient alpha and the internal structure of tests. Psychometrika. 1951;16:297-334'},{id:"B12",body:'Tavakol M, Dennick R. Making sense of Cronbach’s alpha. International Journal of Medical Education. 2011;2:53-55. DOI: 10.5116/ijme.4dfb.8dfd'},{id:"B13",body:'Babbie E. The Practice of Social Research. 12th ed. Belmont, CA: Wadsworth; 2010'},{id:"B14",body:'Polit DF, Beck CT. Generating and Assessing Evidence for Nursing Practice. 8th ed. Williams and Wilkins: Lippincott; 2008'}],footnotes:[],contributors:[{corresp:"yes",contributorFullName:"Kassu Jilcha Sileyew",address:"jkassu@gmail.com",affiliation:'
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