Biomedical subjects

R J Rodgers

Publications and source records attributed to R J Rodgers.

At least 55 records · Page 3Linked to original sources

PubMed1999

Ultrastructure and composition of Call-Exner bodies in bovine follicles.

Call-Exner bodies are present in ovarian follicles of a range of species including human and rabbit, and in a range of human ovarian tumors. We have also found structures resembling Call-Exner bodies in bovine preantral and small antral follicles. Hematoxylin and eosin staining of single sections of bovine ovaries has shown that 30% of preantral follicles with more than one layer of granulosa cells and 45% of small (less than 650 microns) antral follicles have at least one Call-Exner body composed of a spherical eosinophilic region surrounded by a rosette of granulosa cells. Alcian blue stains the spherical eosinophilic region of the Call-Exner bodies. Electron microscopy has demonstrated that some Call-Exner bodies contain large aggregates of convoluted basal lamina, whereas others also contain regions of unassembled basal-lamina-like material. Individual chains of the basal lamina components type IV collagen (alpha 1 to alpha 5) and laminin (alpha 1, beta 2 and delta 1) have been immunolocalized to Call-Exner bodies in sections of fresh-frozen ovaries. Bovine Call-Exner bodies are presumably analogous to Call-Exner bodies in other species but are predominantly found in preantral and small antral follicles, rather than large antral follicles. With follicular development, the basal laminae of Call-Exner bodies change in their apparent ratio of type IV collagen to laminin, similar to changes observed in the follicular basal lamina, suggesting that these structures have a common cellular origin.

Animals↗

PubMed1999

Behavioral effects of diazepam in the murine plus-maze: flumazenil antagonism of enhanced head dipping but not the disinhibition of open-arm avoidance.

Although it is widely believed that benzodiazepines reduce anxiety through positive allosteric modulation of the GABA(A)-chloride channel complex, this is not the only mechanism through which agents of this class can modify CNS function. Furthermore, a significant number of reports of apparent flumazenil blockade of diazepam anxiolysis in animal models have paid limited attention to possible intrinsic behavioral actions of the antagonist per se. In the present study, ethological methods were employed to assess in detail the effects of diazepam, flumazenil, and their combination on the behavior of male DBA/2 mice in the elevated plus-maze paradigm. In two experiments, diazepam (1.5 mg/kg) alone reduced open-arm avoidance and increased head dipping, whereas flumazenil (10-40 mg/kg) alone was without significant behavioral effect. However, with the sole exception of head dipping, prior administration of flumazenil (10 and 40 mg/kg) failed to block the behavioral effects of diazepam under present test conditions. These findings imply that the anxiolytic effects of diazepam in the mouse plus-maze are not mediated through flumazenil-sensitive benzodiazepine receptors and that alternate mechanisms must be considered.

Animals↗

PubMed1999

Influence of spatial and temporal manipulations on the anxiolytic efficacy of chlordiazepoxide in mice previously exposed to the elevated plus-maze.

It has been widely reported that the anxiolytic efficacy of benzodiazepines in the elevated plus-maze test is abolished in subjects (rats or mice) that have been given a single prior undrugged experience of the test apparatus. The present series of experiments was designed to further characterise the key experiential determinants of this intriguing phenomenon in Swiss Webster mice. Using a standard 5 min test duration for both trials, Experiment 1 confirmed the anxiolytic efficacy of chlordiazepoxide (CDP; 5-20 mg/kg) in mice naive to the plus-maze, but a virtual elimination of drug effects in animals that had been pre-exposed to the maze 24 h earlier. Experiments 2 and 3 demonstrated that, while extending the duration of initial exposure to 10 min did not prevent the loss of CDP (10 mg/kg) efficacy in a standard-duration second trial, increasing the duration of both trials reinstated an anxiolytic profile for the compound. Finally, although trial 1 confinement to an open arm did not compromise CDP efficacy when animals were subsequently allowed to freely explore the maze (Experiment 4), closed arm confinement during initial exposure abolished the drug's anxiolytic action upon retest (Experiment 5). In contrast to previous findings in rats, these data suggest that the experientially induced loss of benzodiazepine efficacy in the mouse plus-maze depends rather critically upon prior discovery and exploration of relatively safe areas of the maze (i.e. closed arms). Results are discussed in relation to the hypothesis that the absence of an anxiolytic response to benzodiazepines in plus-maze-experienced subjects reflects the acquisition of an open arm phobia during trial 1.

Animals↗

PubMed1999

Development of the membrana granulosa of bovine antral follicles: structure, location of mitosis and pyknosis, and immunolocalization of involucrin and vimentin.

The membrana granulosa of the ovarian follicle is termed the 'follicular epithelium', yet there have been no studies considering its epithelial nature and how it changes during follicular development. Therefore, these issues were investigated using histology (n = 45 ovaries), considering its structure and the location of proliferating and dying cells, and drawing analogies with other epithelia. Additionally, differences between the layers of granulosa cells were demonstrated by immunohistochemistry (n = 7 ovaries). The structure of the membrana granulosa differed between follicles. Six arbitrary classifications were designed based on these structures, 80 follicles were allocated (n = 13 ovaries) to these classes and the follicular diameters were then measured. For the first time, differences in membrana granulosa structure were shown to correspond to follicle size. Follicles in classes 1-3, where basal granulosa cells were columnar with nuclei positioned basally in the cell, were all < or = 3 mm in diameter. All follicles larger than 3 mm had either columnar basal cells with nuclei positioned centrally (class 4), or had rounded basal cells (class 5), and all follicles > 5 mm had only rounded basal cells. In all these classes, cells in the middle zone were rounded; cells aligning the antrum were often flattened. Irrespective of follicle class, cell proliferation and cell death were shown to be predominantly in the middle portions, rather than the most antral or most basal portions, of the membrana granulosa of healthy and atretic follicles. Involucrin, a marker of keratinocyte differentiation, was localized to the suprabasal region of the membrana granulosa of healthy follicles, particularly in the second and third cellular layers in from the follicular basal lamina. Conversely, the staining intensity for the intermediate filament protein vimentin was lowest in this region, and greatest in the more antral and basal regions. In atretic follicles, there was widespread staining for involucrin and vimentin throughout the membrana granulosa. In conclusion, the membrana granulosa is highly structured, and alters with follicular development. Layers in the membrana granulosa can differ in terms of cell shape, and differ in proliferation and gene expression. In the light of the current work, and an associated study, it is proposed that proliferation occurs in the middle layers, and that granulosa cells, then progress basally or antrally, the latter undergoing terminal differentiation.

Animals↗

PubMed1999

Characterization of the expression of the alternative splicing of the ED-A, ED-B and V regions of fibronectin mRNA in bovine ovarian follicles and corpora lutea.

Fibronectin is an extracellular matrix glycoprotein. Alternative splicing of fibronectin mRNA within three specific regions, the extra domains (ED) A and B and the variable (V) or IIICS region, result in the production of different isoforms of fibronectin. These isoforms differentially regulate tissue developmental processes, such as those occurring during follicular and luteal development. The specific isoforms of fibronectin present in follicles and corpora lutea have not been identified. To identify these, primers for reverse transcription polymerase chain reaction (RT-PCR) were designed to the known bovine amino acid sequence of exons flanking the ED-A, ED-B and V regions. PCR products from bovine fetal liver cDNA were determined to be bovine fibronectin by the correct product size and DNA sequence homology to other species, and to the known bovine amino acid sequence. Bovine ovarian follicles (0.5-9 mm diameter) and corpora lutea (cyclic, early to late mid-luteal phase) were shown to express ED-A+, ED-A-, ED-B+, ED-B-, V+ and V fibronectin isoforms, similar to the liver, lung and kidney of fetuses, but generally not of adult animals. Thus follicles and corpora lutea express isoforms of fibronectin usually expressed in developing tissues.

Alternative Splicing↗

PubMed1999

Evidence for ovarian granulosa stem cells: telomerase activity and localization of the telomerase ribonucleic acid component in bovine ovarian follicles.

We have previously postulated that granulosa cells of developing follicles arise from a population of stem cells. Stem cells and cancer cells can divide indefinitely partly because they express telomerase. Telomerase is a ribonucleoprotein enzyme that repairs the ends of telomeres that otherwise shorten progressively upon each successive cell division. In this study we carried out cell cycle analyses and examined telomerase expression to examine our hypothesis. Preantral (60-100 microm) and small (1 mm) follicles, as well as granulosa cells from medium-sized (3 mm) and large (6-8 mm) follicles, were isolated. Cell cycle analyses and expression of Ki-67, a cell cycle-related protein, were undertaken on follicles of each size (n = 3) by flow cytometry; 12% to 16% of granulosa cells in all follicles were in the S phase, and less than 2% were in the G(2)/M phase. Telomerase activity (n = 3) was highest in the small preantral follicles, declining at the 1-mm stage and even further at the 3-mm stage. In situ hybridization histochemistry was carried out on bovine ovaries, and telomerase RNA was detected in the granulosa cells of growing follicles but not primordial follicles. Two major patterns of staining were observed in the membrana granulosa of antral follicles: staining in the middle and antral layers, and staining in the middle and basal layers. No staining was detected in oocytes. Our results strongly support our hypothesis that granulosa cells arise from a population of stem cells.

Animals↗

PubMed1999

Evidence for alternative pathways of granulosa cell death in healthy and slightly atretic bovine antral follicles.

Granulosa cell death is an early feature of atresia; however, there are many apparent contradictions in the literature concerning the mode of granulosa cell death. We have therefore examined this process in bovine healthy and atretic antral follicles, using a variety of established techniques. Light and electron microscopic observations indicated the presence of pyknotic or shrunken nuclei in both the membrana granulosa and the antrum. In the membrana granulosa, these nuclei were frequently crescent shaped and uniformly electron dense and were approximately the same size as healthy nuclei, all of which are typical of early apoptosis. However, these nuclei were within the membranes of a healthy granulosa cell, suggesting that phagocytosis by a neighboring granulosa cell is an unusually early event in the apoptotic pathway of granulosa cells. In the membrana granulosa, pyknotic nuclei stained intensely with hematoxylin but weakly with the DNA-intercalating stain propidium iodide. A percentage of these pyknotic nuclei stained by TUNEL (terminal deoxy-UTP nick end-labeling). However, in the antrum, the pyknotic nuclei and larger globules of DNA stained intensely with both hematoxylin and propidium iodide, but were not TUNEL positive. The comet assay of cell death produced a streak tail of randomly nicked DNA, rather than the plume of low mol wt apoptotic DNA. Globules collected from fresh follicular fluid stained intensely with propidium iodide and were shown by PAGE to contain DNA, the majority of which was high mol wt. In conclusion, granulosa cells within the membrana granulosa die by apoptosis, with phagocytosis by a neighboring cell preceding any potential budding of the nucleus or cell itself. Granulosa cells near the antrum are sloughed off into the antrum, and their death has features more consistent with that of other cell types that undergo death as a result of terminal differentiation.

Animals↗

PubMed1999

Roles of extracellular matrix in follicular development.

The cellular biology and changes in the extracellular matrix of ovarian follicles during their development are reviewed. During growth of the bovine ovarian follicle the follicular basal lamina doubles 19 times in surface area. It changes in composition, having collagen IV alpha 1-26 and laminin alpha 1, beta 2 and gamma 1 at the primordial stage, and collagen IV alpha 1 and alpha 2, reduced amounts of alpha 3-alpha 5, and a higher content of laminin alpha 1, beta 2 and gamma 1 at the antral stage. In atretic antral follicles laminin alpha 2 was also detected. The follicular epithelium also changes from one layer to many layers during follicular growth. It is clear that not all granulosal cells have equal potential to divide, and we have evidence that the granulosal cells arise from a population of stem cells. This finding has important ramifications and supports the concept that different follicular growth factors can act on different subsets of granulosal cells. In antral follicles, the replication of cells occurs in the middle layers of the membrana granulosa, with older granulosal cells towards the antrum and towards the basal lamina. The basal cells in the membrana granulosa have also been observed to vary in shape between follicies. In smaller antral follicles, they were either columnar or rounded, and in follicles > 5 mm the cells were all rounded. The reasons for these changes in matrix and cell shapes are discussed in relation to follicular development.

Animals↗

PubMed1998

Comparative effects of novel 5-HT1A receptor ligands, LY293284, LY315712 and LY297996, on plus-maze anxiety in mice.

In contrast to the variable efficacy of 5-HT1A receptor full and partial agonists in animal models of anxiety, recent findings in our laboratory have revealed remarkably consistent anxiolytic-like effects for 5-HT1A receptor antagonists in the murine elevated plus-maze paradigm. In the present study, ethological techniques were used directly to compare the plus-maze profiles of three novel ligands varying in intrinsic efficacy at 5-HT1A receptors: LY293284 (full agonist; 0.01-0.3 mg/kg), LY315712 (partial agonist; 0.3-3.0 mg/kg), and LY297996 (antagonist; 0.03-10.0 mg/kg). At the lowest dose tested, LY293284 tended to enhance several indices of anxiety, whereas higher doses suppressed all active behaviours. Although few behavioural effects were observed with LY315712 under present test conditions, a selective reduction in risk assessment was apparent at 1.0-3.0 mg/kg. In contrast to these profiles, LY297996 (3.0-10.0 mg/kg) produced robust anxiolytic-like effects on conventional and ethological parameters but, importantly, did not alter general activity levels. These results provide further support for the anxiolytic potential of 5-HT1A receptor antagonists and are discussed in relation to possible factors underlying inter-laboratory variation in the effects of these agents in animal models of anxiety.

Analysis of Variance↗

PubMed1998

Interindividual variability in Swiss male mice: relationship between social factors, aggression, and anxiety.

In the present study we carried out a series of experiments in Swiss albino male mice to investigate a) the effects of previous social experience on the levels of anxiety in the elevated plus-maze (EPM) and b) whether the response of males in the EPM differs in relation to the different social status. In Experiment 1 we tested in the EPM male mice that received different social experience. Results showed that individually housing generally increased measures of anxiety in the EPM compared with the group-housing condition. Moreover, aggressive males, screened during dyadic encounters in a neutral cage, displayed the highest levels of anxiety relative to the other experimental conditions. In Experiment 2 male mice remained group-housed and were observed to record their social status. Results showed that those animals rated as socially dominant displayed a higher level of EPM anxiety relative to subordinates. From an ethological perspective our findings may be interpreted in terms of coping strategies, with aggressive/dominant animals typified by higher levels of risk assessment and open-arm avoidance than defensive/subordinate animals.

Aggression↗

PubMed1998

Behaviorally selective effects of neuroactive steroids on plus-maze anxiety in mice.

A number of A-ring-reduced metabolites of deoxycorticosterone and progesterone, known to exert agonist activity at the GABA(A) receptor complex, have been reported to reduce anxiety-related behavior in rodents. In the present study, the behavioral selectivity of these effects was assessed in an ethological version of the mouse elevated plus-maze paradigm. Anxiolytic-like profiles, characterised principally by reductions in open arm avoidance measures, were observed following systemic treatment with 5alpha-pregnan-3alpha, 21-diol-20-one (5alpha,3alpha-THDOC; 5.0 and 20.0 mg/kg), 5beta-pregnan-3,20-dione (5beta-DHP; 10-20 mg/kg), 5beta-pregnan-3alpha-ol-20-one (pregnanolone; 20 mg/kg), and 5alpha-pregnan-3alpha-ol-20-one (allopregnanolone; 10-20 mg/kg). In contrast, 5alpha-pregnan-3,20-dione (5alpha-DHP; 10.0-20.0 mg/kg) and 5alpha-pregnan-3beta-ol-20-one (2.5-20.0 mg/kg) were without effect under present test conditions. Detailed behavioral analysis further showed that the antianxiety effects of 5alpha,3alpha-THDOC, 5alpha-DHP, pregnanolone and allopregnanolone were not associated with changes in general activity levels. In addition, profile comparisons revealed that the anxiolytic steroids tend to produce a narrower range of behavioral effects than diazepam (1.0 mg/kg) and, in particular, do not reliably decrease measures of risk assessment. It is concluded that neuroactive steroids produce anxioselective effects in the mouse plus-maze and that their profile of action can at least partially be distinguished from that of a well-characterised benzodiazepine.

5-alpha-Dihydroprogesterone↗

PubMed1998

Responses of Swiss-Webster mice to repeated plus-maze experience: further evidence for a qualitative shift in emotional state?

Behavioral, endocrinological, and pharmacological data suggest that the emotional response of rodents to the elevated plus-maze alters as a function of prior test experience. In the present study, 74 intact male Swiss-Webster mice were exposed to the plus-maze for 5 min on each of 3 consecutive days, with all test sessions recorded on videotape. Behavior patterns for each trial were scored using ethological analysis software and the resultant database subjected to a number of statistical treatments. Analysis of full session profiles (i.e., 5 min total scores) showed that a single prior undrugged experience of the maze increases behavioral indices of anxiety and that these alterations are either maintained or further enhanced on subsequent trials. Furthermore, the behavioral profile evident by trial 3 was largely unchanged when animals were reexposed to the maze 10 days later. More detailed (i.e., min by min) examination of behavior patterns within and between trials demonstrated that unambiguous open arm avoidance is acquired by the third minute of trial 1, and that the behavioral profile evident by the end of trial 1 is (a) markedly different to that seen at the beginning of that trial, and (b) generally maintained or even accentuated on trials 2 and 3. The implied impact of prior test experience on future behavioral strategy in the maze was strongly supported by a series of factor analyses. Thus, while the factor associations of vertical activity and directed exploration remained constant across trials, trial 2 and 3 anxiety measures loaded on a separate factor to that loading trial 1 anxiety measures. A similar trial 1 vs. trials 2 and 3 dissociation was observed for measures of locomotor activity. Although the present findings are consistent with the proposal that prior test experience produces a qualitative shift in emotional response to the elevated plus-maze, the precise basis for this change as well as its full significance for our understanding of anxiety-related processes remain to be determined.

Animals↗

PubMed1998

Benzodiazepine and serotonergic modulation of antipredator and conspecific defense.

The mammalian defense repertory comprises an array of individual behaviors that are extraordinarily sensitive to relevant features of the threat stimulus and the situation in which it occurs. In parallel with increasing awareness of the specificity and complexity of defensive behaviors and of their potential relevance to psychopathologies (e.g. anxiety, panic, and depression) is an escalating use of natural threat stimuli such as attacking conspecifics or predators in paradigms aimed at evaluating drug effects on defense. A review of the literature on benzodiazepine (BZ) and serotonin (5-HT) effects on conspecific and antipredator defense, including defensive analgesia, indicates that both types of stimuli elicit a wide array of relevant defensive behaviors. These studies suggest specificity of drug effects on particular behaviors, rather than a general alteration of all aspects of defense. However, stimulus variability and possible confounding of effects are a considerable problem with conspecific defense paradigms, while antipredator paradigms utilizing human experimenters as the predator may be difficult to use with the domesticated laboratory animal subjects. In addition, sensitivity to the organization of defensive behaviors and to differences between species in defense patterns is necessary to adequate interpretation of results. Nonetheless, these paradigms have permitted major advancements in analysis of the behavioral defense systems and their sensitive use in drug studies will greatly facilitate an understanding of the physiology of defense.

Agonistic Behavior↗

PubMed1998

Tolerance to acute anxiolysis but no withdrawal anxiogenesis in mice treated chronically with 5-HT1A receptor antagonist, WAY 100635.

Anxiolytic-like activity in the mouse elevated plus-maze has recently been demonstrated for a range of compounds varying in degree of selectivity as 5-HT1A receptor antagonists. As tolerance and dependence liability are among the major clinical disadvantages of benzodiazepine therapy, the present study examined the effects of acute drug challenge on the plus-maze profiles of mice following daily treatment for 20 days with saline, chlordiazepoxide (CDP; 10.0 mg/kg) or the selective 5-HT1A receptor antagonist, WAY 100635 (0.1-1.0 mg/kg). To assess the development of physical dependence (withdrawal anxiogenesis), the study incorporated independent groups of animals tested on the maze 24 h after the final dose. Challenge with CDP or WAY 100635 produced behavioural changes indicative of anxiety reduction in mice that had received daily handling/saline for 20 days, thereby demonstrating that the chronic injection regimen per se had not compromised the acute efficacy of either agent. The absence of a similar response to acute drug challenge in mice treated chronically with CDP or WAY 100635 suggested the development of tolerance to the acute anxiolytic effects of both compounds under present test conditions. Despite these observations, however, no signs of enhanced anxiety were evident 24 h following discontinuation of chronic treatment with either compound. In a further experiment, the absence of withdrawal anxiogenesis at 24 h was replicated and extended to discontinuation periods of 36 and 48 h for both drugs. Although present results show that tolerance develops to the acute anxiolytic effects of CDP and WAY 100635 in the murine plus-maze, they also suggest that enhanced anxiety is not an inevitable consequence of abrupt cessation of chronic treatment with either compound.

Animals↗

PubMed1998

Changes in testicular steroidogenic acute regulatory (STAR) protein, steroidogenic enzymes and testicular morphology associated with increased testosterone secretion in bulls receiving the luteinizing hormone releasing hormone agonist deslorelin.

Testosterone secretion and the expression and relative contents of steroidogenic acute regulatory (StAR) protein and steroidogenic enzymes cholesterol side-chain cleavage cytochrome P450 (P450SCC), 3beta-hydroxysteroid dehydrogenase/delta(5)-->delta(4)-isomerase (3 beta-HSD), and (17)alpha-hydroxylase cytochrome P450/C17-20 lyase (P450(17)alpha) were determined in testicular tissues of bulls treated with a LHRH agonist. Testis morphology and spermatogenesis were also examined. In Experiment 1, bulls (30-mo-old) received no treatment (control, n = 7) or were implanted for 10 days with the LHRH agonist deslorelin (n = 7). Bulls were castrated on Day 10 and testis tissues prepared for Western and Northern blotting. At castration, bulls implanted with deslorelin had greater plasma testosterone (5-fold) and testis content of testosterone (10-fold) compared with control bulls. Relative content (per micrograms total testis protein or RNA) of StAR protein, 3beta-HSD, P450SCC, and mRNA for P450(17)alpha in bulls treated with deslorelin ranged from 3- to 6-fold that of control bulls. In Experiment 2, bulls (20-mo-old) were left untreated (control, n = 6) or implanted with deslorelin (n = 12) for 120 days. On Day 120, bulls were castrated and right testis tissues prepared for morphology. Testis volume and weight were increased (P < 0.01) in bulls treated with deslorelin compared with control bulls. Stereological analysis revealed that this increase occurred in all compartments (seminiferous epithelium, lumen and interstitium) studied, but was significant (P < 0.01) only for the seminiferous epithelium. Absolute numbers of round spermatids per testis were increased (P < 0.05) in bulls treated with deslorelin compared with control bulls. Increased testosterone secretion in bulls treated with deslorelin was associated with increased testicular StAR protein and steroidogenic enzymes. Bulls treated long-term with deslorelin had a faster rate of testis growth and increased daily sperm production at the end of the experiment.

17-Hydroxysteroid Dehydrogenases↗

PubMed1998

Distribution of the alpha1 to alpha6 chains of type IV collagen in bovine follicles.

During follicular development the proliferative and differentiated state of the epithelioid granulosa cells changes, and the movement of fluid across the follicular basal lamina enables the formation of an antrum. Type IV collagen is an important component of many basal laminae. Each molecule is composed of three alpha chains; however, six different type IV collagen chains have been identified. It is not known which of these chains are present in the follicular basal lamina and whether the type IV collagen composition of the basal lamina changes during follicular development. Therefore, we immunolocalized each of the six chains in bovine ovaries using antibodies directed to the nonconserved non-collagenous (NC) domains. Additionally, dissected follicles were digested with collagenase to release the NC domains, and the NC1 domains were then detected by standard Western immunoblot methods. The follicular basal lamina of almost all primordial and preantral follicles was positive for all type IV collagen alpha chains. Colocalization of type IV collagen and factor VIII-related antigen allowed for discrimination between the follicular and endothelial basal laminae. Type IV collagen alpha1, alpha2, alpha3, alpha4, and alpha5 chains were present within the follicular basal lamina of only a proportion of antral follicles (17 of 22, 20 of 21, 15 of 18, 14 of 28, and 12 of 23, respectively), and staining was less intense than in the preantral follicles. Staining for the alpha1 and alpha2 chains was diffusely distributed throughout the theca in regions not associated with recognized basal laminae. The specificity of this immunostaining for alpha1 and alpha2 chains of type IV collagen was confirmed by Western immunoblots. As well as being detected in the basal lamina of approximately half of the antral follicles examined, type IV collagen alpha4 also colocalized with 3beta-hydroxysteroid dehydrogenase-immunopositive cells in the theca interna. Type IV collagen alpha6 was detected in the basal lamina of only one of the 16 antral follicles examined. Thus, the follicular basal lamina changes in composition during follicular development, with immunostaining levels being reduced for all type IV collagen chains and immunoreactivity for type IV collagen alpha6 being lost as follicle size increases. Additionally, immunoreactivity for alpha1 and alpha2 appears in the extracellular matrix of the theca as it develops.

3-Hydroxysteroid Dehydrogenases↗

PubMed1998

Expression of types 1, 2, and 3 17 beta-hydroxysteroid dehydrogenase in subcutaneous abdominal and intra-abdominal adipose tissue of women.

Human adipose tissue is known to have 17 beta-oxidoreductase activity, interconverting estrone (E1) and estradiol (E2), as well as androstenedione (A) and testosterone (T). We examined both the subcutaneous abdominal and intra-abdominal (visceral) adipose tissue of women for expression of types 1, 2, and 3 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) using ribonuclease (RNase) protection assay and RT-PCR/Southern blotting. Type 1 17 beta-HSD, which encodes the enzyme responsible for the conversion of E1 to E2 in the placenta and ovary, was expressed in the subcutaneous abdominal and intra-abdominal adipose tissue of women, but the messenger RNA transcripts were predominantly incompletely spliced and therefore unlikely to encode an active protein. A pseudogene for type 1 17 beta-HSD was also expressed in these tissues, but messenger RNA transcripts were again unspliced. Type 2 17 beta-HSD, which encodes an enzyme that can catalyze the conversion of T to A and E2 to E1, was expressed in both the subcutaneous abdominal and intra-abdominal adipose tissue of women. Type 3 17 beta-HSD was also expressed in adipose tissue from both sites studied. Type 3 17 beta-HSD encodes the enzyme that catalyzes the conversion of A to T in the testis and also converts E1 to E2. Together with aromatase, which is known to be expressed in adipose tissue, the expression of types 2 and 3 17 beta-HSD indicates that sex steroid production in the adipose tissue of women is a complex process. The association of visceral obesity with the development of insulin resistance and dyslipidaemia raises the question of the role of steroid production in adipose tissue in the pathogenesis of these disorders.

17-Hydroxysteroid Dehydrogenases↗

PubMed1998

Differential localization of laminin chains in bovine follicles.

The composition of a basal lamina markedly affects its ability to filter material and affects the fate of adjacent epithelial cells. Therefore, basal laminae differ in composition with tissue development, and between different tissues in the body. Laminins are a component of basal laminae and consist of one alpha, one beta and one gamma chain, of which there are at least five, three and two isoforms, respectively. This is the first study to immunolocalize a range of these individual laminin chains (alpha 1, alpha 2, beta 1, beta 2, gamma 1) in ovarian follicles. Frozen sections of bovine ovaries (n = 6) were immunostained using specific antisera to laminin chains and factor VIII-related antigen (to identify endothelial cells). Secondary antisera were labelled with one of two different fluorochromes (DTAF and Cy3), and dual localization of laminin chains and factor VIII-related antigen was performed. The alpha 1, beta 2 and gamma 1 chains were consistently localized to the follicular basal lamina in all healthy follicles. Staining was less intense in the atretic antral follicles. Conversely, alpha 2 and beta 1 were rarely present in the follicular basal laminae of healthy antral follicles. Two of nine healthy antral follicles observed stained weakly for alpha 2 in their basal lamina, and beta 1 was present at low concentrations in growing preantral follicles. In atretic antral follicles, the follicular basal lamina stained positively for alpha 1, alpha 2, and beta 2 but no beta 1 was detected and the gamma 1 staining was less intense than in healthy follicles. Antisera to Englebreth Holm-Swarm tumour laminin stained basal laminae of all follicles. In the theca of antral follicles, beta 1 and beta 2 chains were both present in the vasculature. Staining for the gamma 1 chain was present in the thecal vasculature and generally throughout the theca of healthy and atretic antral follicles. Therefore, the composition of the follicular basal lamina alters during development and atresia, and potentially plays a role in the changing identity of the granulosa cells and the accumulation of antral follicular fluid.

Animals↗