Testicular Sertoli cells make a niche for the division and differentiation of germ cells. Sertoli cells respond to increased follicle-stimulating hormone (FSH) and testosterone (T) levels at the onset of puberty by producing paracrine factors which affect germ cells and trigger robust onset of spermatogenesis. Such paracrine support to germ cells is absent during infancy, despite Sertoli cells being exposed to high FSH and T within the infant testis. This situation is similar to certain cases of male idiopathic infertility where post-pubertal Sertoli cells fail to support germ cell division and differentiation in spite of endogenous or exogenous hormonal support. Defective Sertoli cells in such individuals may fail to express the full complement of their paracrine repertoire. Identification and supplementation with such factors may overcome Sertoli cells deficiencies and help trigger quantitatively and qualitatively normal differentiation of germ cells. To this end, we compared the transcriptome of FSH- and T-treated infant and pubertal monkey Sertoli cells by DNA microarray. Expression of Wnt3, a morphogen of the Wnt/β-catenin pathway, was higher in pubertal Sertoli cells relative to infant Sertoli cells. Transgenic mice were generated by us in which Wnt3 expression was curtailed specifically in post-pubertal Sertoli cells by shRNA. Subfertility and oligozoospermia were noticed in such animals with low Wnt3 expression in post-pubertal Sertoli cells along with diminished expression of Connexin43, a gap-junctional molecule essential for germ cell development. We report that the FSH- and T-targetedf Wnt3 governs Sertoli cell-mediated regulation of spermatogenesis and hence is crucial for fertility.
Bacteria belonging to the Alphaproteobacteria normally harbour multiple copies of the heat shock sigma factor (known as s 32, s H or RpoH). Azospirillum brasilense, a non-photosynthetic rhizobacterium, harbours five copies of rpoH genes, one of which is an rpoH2 homologue. The genes around the rpoH2 locus in A. brasilense show synteny with that found in rhizobia. The rpoH2 of A. brasilense was able to complement the temperature-sensitive phenotype of the Escherichia coli rpoH mutant. Inactivation of rpoH2 in A. brasilense results in increased sensitivity to methylene blue and to triphenyl tetrazolium chloride (TTC). Exposure of A. brasilense to TTC and the singlet oxygen-generating agent methylene blue induced several-fold higher expression of rpoH2. Comparison of the proteome of A. brasilense with its rpoH2 deletion mutant and with an A. brasilense strain overexpressing rpoH2 revealed chaperone GroEL, elongation factors (Ef-Tu and EF-G), peptidyl prolyl isomerase, and peptide methionine sulfoxide reductase as the major proteins whose expression was controlled by RpoH2. Here, we show that the RpoH2 sigma factor-controlled photooxidative stress response in A. brasilense is similar to that in the photosynthetic bacterium Rhodobacter sphaeroides, but that RpoH2 is not involved in the detoxification of methylglyoxal in A. brasilense.
Background: Abnormal uterine bleeding (AUB) is a common gynecological problem associated with considerable morbidity and significantly affects the patients. The aim of the study was to analyze the histopathological patterns of endometrium in patients presenting with AUB and also to determine the incidence of AUB in various age groups.Methods: This is a retrospective study, conducted in the Department of Obstetrics and Gynaecology, in a tertiary care teaching hospital, Mumbai, India from March 2016 till date. All cases of AUB were included in the study. Data was entered in microsoft excel and managed in statistical package for the social sciences (SPSS) version 16. Analysis was done in the form of percentages and proportions and represented as tables where necessary.Results: A total of 120 cases were analyzed. Patients’ age ranged from 22-79 years. AUB was most common among the perimenopausal females (41-50years). The most common presenting symptom was heavy menstrual bleeding (53%). Dilatation and curettage (D&C) was performed in all cases and 96 underwent hysterectomy as final resort. Endometrial proliferative pattern was the most common histopathological finding and was seen in 27% patients, followed by endometrial hyperplasia in 13.5% patients, secretory endometrium (12.7%) and disordered proliferative endometrium were seen in 10.9% patients each. Malignancy was detected in 1.7% of cases and endometrial carcinoma was the most common lesion.Conclusions: Endometrial sampling is especially indicated in women above the age of 35 years to rule out malignancy and preneoplasia. Among the females with no organic pathology, normal physiological patterns with proliferative, secretory, and menstrual changes were observed. The most common endometrial pathology in this study was endometrial proliferation.
In response to various hormonal (follicle-stimulating hormone [FSH] and testosterone [T]) and biochemical inputs, testicular Sertoli cells (Sc) produce factors that regulate spermatogenesis. A number of FSH- and T-responsive Sc-specific genes, necessary for spermatogenesis, have been identified to date. However, the hormone-induced in vitro expression pattern of most of these genes is reported to be inconsistent at various time points in primary rat Sc cultures. As a matter of convenience, cultured Sc are constantly exposed to hormones for a few hours to days in the reported literature, although Sc are exposed to pulsatile FSH and T in vivo. The major aim of the present study is to evaluate the advantage, if any, of the in vitro administration of pulsatile hormone (FSH and T in combination) treatment on gene expression of cultured Sc as compared with that of constant hormone treatment. Pulsatile treatment (a 30-min hormonal exposure every 3 h) mimicking the in vivo condition reveals a more prominent effect of hormones in augmenting gene expression as compared with constant treatment. Our results indicate that the expressions of Stem cell factor (Scf, only responsive to FSH), Claudin11 (only responsive to T) and Transferrin (both FSH- and T-responsive) mRNAs are significantly higher at 12 h upon pulsatile treatment than upon constant hormonal treatment. Maximal expression of relevant genes because of pulsatile treatment with hormones suggests that this protocol provides a more suitable premise for assessing hormone-induced gene expression in isolated Sc than one involving constant exposure to hormones.
Our ability to decipher gene sequences has increased enormously with the advent of modern sequencing tools, but the ability to divulge functions of new genes have not increased correspondingly. This has caused a remarkable delay in functional interpretation of several newly found genes in tissue and age specific manner, limiting the pace of biological research. This is mainly due to lack of advancements in methodological tools for transgenesis. Predominantly practiced method of transgenesis by pronuclear DNA-microinjection is time consuming, tedious, and requires highly skilled persons for embryo-manipulation. Testicular electroporation mediated transgenesis requires use of electric current to testis. To this end, we have now developed an innovative technique for making transgenic mice by giving hypotonic shock to male germ cells for the gene delivery. Desired transgene was suspended in hypotonic Tris-HCl solution (pH 7.0) and simply injected in testis. This resulted in internalization of the transgene in dividing germ-cells residing at basal compartment of tubules leading to its integration in native genome of mice. Such males generated transgenic progeny by natural mating. Several transgenic animals can be generated with minimum skill within short span of time by this easily adaptable novel technique.
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