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1.
Histol Histopathol ; : 18660, 2023 Sep 04.
Article in English | MEDLINE | ID: mdl-37721417

ABSTRACT

There is increasing interest in understanding the tissue biology of human amniotic membrane (hAM) given its applications in medicine. One cellular component is mesenchymal cells, which can be extracted, cultured and differentiated "in vitro" into various cell types. These studies show that there is heterogeneity among mesenchymal cells. The aim of this work is to study the membrane "in situ" to determine whether this cellular heterogeneity exists. The hAMs were obtained from caesarean deliveries at term and analyzed by histological techniques. Types I-III mesenchymal cells and Hofbauer were distinguished by light microscopy. Histochemically, mesenchymal cell types showed successively increasing positivity to: PAS, vimentin, fibronectin, and Concanavalin-A; VGEF, TGF-ß2, PDGF-C, FGF-2. By the semiquantitative point of view, the percentage of Type II cells was 60%, significantly higher than the other types. With transmission electron microscopy, an intermediate cell type between II-III was observed. Strong vesiculation of the rough endoplasmic reticulum (RER) with exocytosis was observed. In addition, an accumulation of a similar material to the extracellular matrix in the RER caused its dilation especially in type III cells. Some of this material acquired a globular structure. These structures were also found free in the extracellular matrix. In conclusion, the mesenchymal cells of the fibroblastic layer of the hAMs studied are heterogeneous, with some undifferentiated and others with a probably senescent fibroblastic phenotype with accumulation in their RER of fibronectin. These results may be of interest to extract mesenchymal cells from hAMs for use in regenerative medicine and to better understand the mechanisms of fetal membrane rupture.

2.
Sci Rep ; 10(1): 13360, 2020 08 07.
Article in English | MEDLINE | ID: mdl-32770046

ABSTRACT

Granulocyte-macrophage colony stimulating factor (GM-CSF) is a pro-inflammatory cytokine identified in boar seminal plasma (SP) but until now unexplored in terms of place of production and its association to spermatozoa. This study aimed to explore these aspects by evaluating the presence of GM-CSF in porcine reproductive organs (testes, epididymis and accessory sex glands), SP and mature spermatozoa (from cauda epididymis and ejaculated) using Western blot (WB), immunohistochemistry and immunocytochemistry. Positive labelling was obtained in tissues, SP and spermatozoa. In reproductive organs, WB revealed three forms of GM-CSF with different glycosylation degrees (15, 31 and 40 kDa). In SP and epididymal fluid, the GM-CSF appeared only in its active form while in spermatozoa the GM-CSF form present varied among sperm sources. Non-viable spermatozoa showed more GM-CSF than viable spermatozoa (14.87 ± 1.98 RU vs. 7.25 ± 0.52 RU) of fluorescence intensity. In conclusion, GM-CSF is widely present in the reproductive tract of male pigs, attached to the spermatozoa already in the epididymis as well as verted to SP. Consequently, the GM-CSF ought to regulate male genital tract and sperm function as well as mediating initial inflammatory responses and further mediating later immune actions by the female to semen deposition.


Subject(s)
Genitalia, Male/metabolism , Granulocyte-Macrophage Colony-Stimulating Factor/metabolism , Semen/metabolism , Spermatozoa/metabolism , Animals , Epididymis/metabolism , Male , Swine , Testis/metabolism
3.
Mol Reprod Dev ; 75(9): 1473-83, 2008 Sep.
Article in English | MEDLINE | ID: mdl-18288675

ABSTRACT

The glycoside residues (glycoconjugates, GC) of the zona pellucida (ZP) glycoproteins are important during the first phases of fecundation. Our aim in this work was to determine the lectin affinity pattern of porcine ZP in order to analyze the changes that take place during: (a) preantral folliculogenesis, (b) the follicular atresia process, and (c) antral growth. Several prepubertal and adult pig ovaries and different sized antral follicles were used. Conventional carbohydrate histochemical techniques and peroxidase and digoxigenin (DIG) lectins were used to reveal the acid groups and the glycosidic residues of the ZP. It was seen that the ZP forms in the preantral follicles throughout their growth period. In primordial and primary follicles, ZP in the process of formation showed neutral GC. SBA, RCA-I, MAA, WGA lectins, and AAA after methylation-saponification (MS) were positive in the ZP of primordial and primary follicles. The affinity for SBA, RCA-I, MAA, and WGA increased in the multilaminar-primary follicles and new affinities for UEA-I and LFA were observed. After MS, AAA, SNA, PNA, and SBA reactivity was observed. The ZP of antral follicle oocytes of different sizes showed the same lectin pattern as multilaminar-primary follicles. The oocyte ooplasm and the follicular fluid of large antral follicles showed less affinity for WGA and LFA lectins and less intensive staining with AB (pH 2.5). Atresia did not change the antral or preantral follicle oocyte ZP lectin pattern. In conclusion, the follicles showed substantial changes in their ZP glycosidic composition as they developed, especially, during the change from primary to multilaminar-primary follicles. The ZP glycosidic composition showed no significant change during the growth of antral follicles and follicular atresia in our study.


Subject(s)
Cytoplasm/chemistry , Follicular Atresia/physiology , Glycosides/analysis , Oocytes/chemistry , Ovarian Follicle/growth & development , Sus scrofa , Zona Pellucida/chemistry , Animals , Cell Size , Female , Histocytochemistry , Lectins/analysis , Lectins/chemistry , Lectins/metabolism , Oocytes/metabolism , Ovarian Follicle/cytology , Ovarian Follicle/physiology , Sus scrofa/physiology , Zona Pellucida/metabolism
4.
J Androl ; 28(1): 123-35, 2007.
Article in English | MEDLINE | ID: mdl-16957139

ABSTRACT

Aging and short photoperiod exposure induce germ cell apoptosis in the Syrian hamster; however, the specific germ cells affected and the molecular pathways triggered have not been elucidated. We analyzed germ cell apoptosis and the expression of the Fas/Fas-L system, Bcl-2 family, and p53 in aged and photoinhibited hamsters and compared with those young maintained in natural photoperiod. Aging increased apoptosis in spermatogonia and spermatocytes, but in photoinhibited hamster testes only an increase in apoptotic spermatocytes was observed. Apoptosis was higher in aged hamsters in stages I-IV, V-VI and VII-VIII. Aging increased apoptosis of spermatogonia in stages I-IV and V-VI. Apoptotic pachytene spermatocytes were significantly higher in stages I-IV, V-VI, and VII-VIII in aging. Apoptotic preleptotene and pachytene spermatocytes were higher in aging, but no differences were observed in leptotene-zygotene. Fas-L was expressed by Sertoli cells, of young, aged, and photoinhibited hamsters. Bcl-x(L) was strongly expressed in germ cells on young hamsters and slightly in aging and after short photoperiod exposure. Spermatocytes of photoinhibited hamsters were intensively stained with Fas, Bax, Bcl-xs/L, and p53. In conclusion, aging increases apoptosis in spermatogonia and spermatocytes, depending on the stage of the seminiferous epithelium cycle, whereas after a short photoperiod exposure only an increase in apoptotic spermatocytes is observed. The results suggest that Fas, Bcl-x(L), Bax, and p53 participate in germ cell apoptosis induction after short photoperiod exposure, whereas only Bcl-x(L) is involved in aging.


Subject(s)
Aging/physiology , Apoptosis , Gene Expression , Photoperiod , Seminiferous Epithelium/pathology , Seminiferous Epithelium/physiology , Spermatozoa/physiology , Aging/metabolism , Animals , Cricetinae , Fas Ligand Protein/genetics , Fas Ligand Protein/physiology , Gene Expression/physiology , Gene Expression/radiation effects , Immunoenzyme Techniques , Male , Mesocricetus , Proto-Oncogene Proteins c-bcl-2/genetics , Proto-Oncogene Proteins c-bcl-2/physiology , Seminiferous Epithelium/metabolism , Spermatozoa/pathology , Spermatozoa/radiation effects , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/physiology , fas Receptor/genetics , fas Receptor/physiology
5.
J Mol Histol ; 36(4): 235-42, 2005 May.
Article in English | MEDLINE | ID: mdl-16200455

ABSTRACT

It has been shown that there are sugars in corpora amylacea, but little attention has been focused on the expression of glycoconjugates in corpora amylacea of normal and hyperplastic prostatic glands. The present study characterizes and compares the expression of glycoconjugates in corpora amylacea of normal and hyperplastic prostatic glands of elderly men by using alcian blue (AB) stain and lectin histochemistry. Corpora amylacea were larger and more numerous in hyperplastic glands compared to normal glands. The stain with AB revealed the presence of sulfated and carboxyl components in corpora amylacea. In hyperplastic prostatic glands the sulfur and acid contents of corpora amylacea were increased. Lectin affinities of corpora amylacea from normal prostatic glands demonstrated the presence of fucose, mannose, sialic acid, N-acetyl galactosamine and N-acetyl glucosamine residues. In the hyperplastic glands the lectin binding pattern of corpora amylacea was qualitatively similar to normal glands, but an increase in GalNAc, sialic acid, mannose and fucose residues was observed. Normal prostatic glands showed a weak to moderate content of mannose residues, and in contrast a strong GNA and Con-A staining was observed in hyperplastic glands. MAA and SNA affinities indicated that the content of sialic acid residues was higher in hyperplastic glands compared with normal prostatic glands. Also NAcGal residues were increased in hyperplastic glands. Luminal secretion, secretory cells and apical border of epithelium showed a similar although more intense Lectin-binding pattern as compared with corpora amylacea both in normal and hyperplastic prostatic glands. Lectin histochemistry shows that the glycoconjugates expressed in the glandular epithelium are similar to those found in corpora amylacea both in normal and hyperplastic glands. In addition, in hyperplastic glands, where the corpora amylacea are higher in size and more numerous, the reaction to lectins is more intense especially with mannose and sialic acid residues. The results suggest that corpora amylacea are originated at least in part from prostatic secretion.


Subject(s)
Glycoconjugates/metabolism , Prostate/metabolism , Prostate/pathology , Humans , Hyperplasia , Immunohistochemistry , Lectins/metabolism , Male , Prostate/anatomy & histology , Prostate/cytology , Substrate Specificity
6.
Acta Histochem ; 107(5): 365-72, 2005.
Article in English | MEDLINE | ID: mdl-16185749

ABSTRACT

Cryptorchidism is a frequent male sexual disorder in mammals, which affects the histology of the tunica propria, interstitial tissue, blood vessels, seminiferous epithelium and testis functioning. In this paper, proliferation and apoptosis were examined in the seminiferous epithelium of both testes from unaffected boars and from boars suffering unilateral and bilateral cryptorchidism. In germ cells, proliferation was studied using the immunohistochemical PCNA technique, and apoptosis was analysed by in situ TUNEL labelling. An index was obtained for the proliferation and apoptosis observed in seminiferous tubules. In abdominal testes the epithelium contained few spermatogonia and Sertoli cells. In the testes of unaffected boars, numerous spermatogonia proliferated, whereas in cryptorchid testes such proliferation was lower and the proliferation/apoptosis ratio diminished. In the unaffected group, the TUNEL-positive germ cells were spermatogonia and spermatocytes in different phases of meiosis. In abdominal testes, the TUNEL-positive germ cells were spermatogonia alone. The apoptosis index of both abdominal and scrotal testes was similar. In conclusion, spontaneous cryptorchid testes showed a lower rate of spermatogonia proliferation in the seminiferous epithelium.


Subject(s)
Apoptosis , Cryptorchidism/pathology , Spermatogonia/pathology , Testis/pathology , Animals , Cell Proliferation , Cryptorchidism/physiopathology , Immunohistochemistry , In Situ Nick-End Labeling , Male , Proliferating Cell Nuclear Antigen/analysis , Seminiferous Epithelium/pathology , Seminiferous Epithelium/physiopathology , Sertoli Cells/pathology , Sertoli Cells/physiology , Sexual Maturation , Spermatocytes/pathology , Spermatocytes/physiology , Spermatogenesis , Spermatogonia/physiology , Sus scrofa , Testis/physiopathology
7.
Acta Histochem ; 105(2): 165-73, 2003.
Article in English | MEDLINE | ID: mdl-12831168

ABSTRACT

The objective of the present study was to characterize glycoconjugates of hamster testis in gonadally-active and -inactive states by lectin histochemical methods. Thirteen HRP- or digoxigenin-labeled lectins were used in samples obtained from fertile and photoinhibited hamsters. In gonadally-active hamsters, spermatozoa tails were stained with Con-A, HPA, PNA, UEA-I, LTA, AAA, WGA and LFA and weakly with GNA and RCA-I. Spermatozoa acrosomes were labeled with HPA, SBA, WGA and PNA. Spermatid acrosomes were labeled with SBA, RCA-I, PNA, and WGA. Staining with GNA and Con-A was found in the Golgi phase and HPA staining was found in the Golgi phase and maturated spermatids. Cytoplasm of spermatocytes was labeled with Con-A, GNA, LTA, AAA, RCA-I, HPA, WGA and LFA, whereas spermatocyte membranes were stained with Con-A, LTA and AAA. Spermatogonia were strongly labeled with Con-A and moderately labeled with AAA, WGA and LFA. Sertoli cells were positive after staining with Con-A, AAA, WGA, and LFA. The lamina propria was positive after staining with UEA-I, LTA, AAA and LFA. Leydig cells showed strong labeling with SBA, Con-A, GNA, SNA and MAA, moderate labeling with WGA, weak labeling with RCA-I, AAA and LFA. In gonadally-inactive hamsters, spermatocytes showed increased staining with HPA, PNA and AAA, whereas staining with Con-A, GNA and LTA had disappeared. Spermatogonia showed an increased labeling with AAA and WGA, but labeling with Con-A and LFA had disappeared. Sertoli cells were strongly labeled with GNA. Con-A and GNA staining was decreased in Leydig cells of gonadally-inactive hamsters but PNA and HPA staining was increased. The lamina propria in regressed testes showed intense labeling with PNA. These results suggest that histological, morphological and hormonal changes occurring in hamster testis during exposure to a short photoperiod are reflected in altered patterns of expression and distribution of N- and O-linked glycans.


Subject(s)
Glycoconjugates/analysis , Photoperiod , Testis/chemistry , Animals , Cricetinae , Histocytochemistry/methods , Lectins/analysis , Leydig Cells/chemistry , Leydig Cells/cytology , Male , Polysaccharides/analysis , Testis/anatomy & histology
8.
Int J Androl ; 25(5): 281-7, 2002 Oct.
Article in English | MEDLINE | ID: mdl-12270025

ABSTRACT

In the hamster, male reproductive quiescence is accomplished via testicular atrophy and the germinal epithelium is regressed to spermatogonia and spermatocytes after 8-14 weeks of short photoperiods. However, the cellular mechanisms involved in this process have not been elucidated. As it is suggested that the regulation of seasonal testicular activity is characterized by coordinated shifts in the relationships between mitosis, meiosis and apoptosis, the changes in the proliferative and apoptotic activity in the seminiferous epithelium of photoinhibited Syrian hamster were examined and compared with those maintained in natural photoperiod. The proliferative activity was studied using BrdU immunostaining, and germ cell apoptosis was assessed by in situ TUNEL labelling and transmission electron microscopy. A significant increase in the rate of apoptosis (percentage of TUNEL-positive spermatogonia + spermatocytes) was observed in photoinhibited animals (2.84 +/- 0.16) compared with those exposed to natural photoperiod (0.77 +/- 0.03, p < 0.05). The majority of apoptotic germ cells were spermatocytes and in some occasions spermatogonia. Germ cell apoptosis was confirmed by morphological characteristics: condensation of the chromatin and nuclear fragmentation. The rate of proliferation (percentage of BrdU-positive spermatogonia + preleptotene spermatocytes) was significantly higher in photoinhibited hamsters (42.7 +/- 2.6) compared with animals exposed to natural photoperiod (31.1 +/- 1.6, p < 0.05). After the exposure to a short photoperiod the apoptotic index positively correlated with the proliferative index (r = 0.8150, p < 0.05). In conclusion, the seminiferous epithelium of photoinhibited Syrian hamsters is characterized by an increased rate of apoptosis associated to an enhanced rate of proliferation.


Subject(s)
Apoptosis , Cell Division , Photoperiod , Seminiferous Tubules/cytology , Animals , Cricetinae , Epithelium/ultrastructure , In Situ Nick-End Labeling , Male , Mesocricetus , Microscopy, Electron, Scanning , Seminiferous Tubules/ultrastructure
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