Stage-specific expression of RESP18 in the testes.

RESP18 (regulated endocrine-specific protein of 18 KD) is an endoplasmic reticulum (ER) protein that was identified by coordinate dopaminergic regulation with pro-opiomelanocortin in the rat neurointermediate pituitary. Many attributes of RESP18 suggest an important function in neuroendocrine cells. Several neuropeptides, growth factors, and enzymes involved in biosynthesis of classical chemical neurotransmitters, have been identified in germ cells, Sertoli cells, and spermatozoa. In this study, screening of reproductive tissues revealed high levels of RESP18 protein and mRNA in the testes but not in ovaries or epididymis. The testes and sperm expressed 18-KD RESP18 and a unique 19-KD isoform. To better understand RESP18 expression in the testes, we have examined the stages of the cycle of the seminiferous epithelium by immunohistochemistry and Western blot analyses. Immunohistochemical analysis showed that RESP18 protein was expressed exclusively in spermatocytes and maturing spermatids. RESP18 protein was expressed at high levels in Step 1-8 round spermatids, in which the PC4 prohormone convertase, nerve growth factor, and proenkephalin are also expressed. Western blots, Northern blots, and indirect immunofluorescence staining demonstrated RESP18 expression in sperm.

Download Full-text

Protein expression pattern of calcium-responsive transactivator in early postnatal and adult testes

Histochemistry and Cell Biology ◽

10.1007/s00418-020-01942-1 ◽

2021 ◽

Author(s):

Ana Du ◽

Li Li ◽

Zhaoshuang Jiao ◽

Gaochun Zhu ◽

Ting Peng ◽

...

Keyword(s):

Epithelial Cells ◽

Protein Expression ◽

Expression Pattern ◽

Sertoli Cells ◽

Immunohistochemical Analysis ◽

The Body ◽

Spermatogenic Cells ◽

Specific Expression ◽

Adult Rat ◽

Regulation Of Cell Cycle

AbstractCalcium-responsive transactivator (CREST), a nuclear protein highly expressed in postmitotic neurons, is involved in the regulation of cell cycle, differentiation and dendritic development of neuronal cells. Its mRNA has been detected in the testis of adult rat, whilst its protein expression and distribution pattern in the testis remain to be elucidated. In this study, we examined the distribution of CREST in the adult testes of both rats and human as well as the expression pattern of CREST in the testes of postnatal developing rats. In the adult testes of both human and rats, immunohistochemical analysis revealed that CREST was selectively distributed in the mature Sertoli cells but not in the spermatogenic cells. In the testes of postnatal developmental rats, CREST was expressed not only in Sertoli cells but also in the gonocytes and spermatogenic cells at the initial stage of spermatogenic cell differentiation. CREST immunoreactivity continued to increase in Sertoli cells during differentiation, reaching its peak in adulthood. However, CREST immunostaining intensity dramatically decreased as the spermatogenic cells differentiate, disappearing in the post-differentiation stage. Furthermore, Brg1 and p300, two CREST-interacting proteins ubiquitously expressed in the body, are found to be colocalized with CREST in the spermatogenic epithelial cells including Sertoli cells. The unique expression pattern of CREST in developing testis suggests that CREST might play regulatory roles in the differentiation of spermatogenic epithelial cells. The Sertoli cell-specific expression of CREST in the adulthood hints that CREST might be a novel biomarker for the mature Sertoli cells.

Download Full-text

Expression of Platelet Membrane Glycoprotein V in Human Megakaryocytes and Megakaryocytic Cell Lines: A Study Using a Novel Monoclonal Antibody against GPV

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648955 ◽

1994 ◽

Vol 72 (05) ◽

pp. 762-769 ◽

Cited By ~ 11

Author(s):

Toshiro Takafuta ◽

Kingo Fujirmura ◽

Hironori Kawano ◽

Masaaki Noda ◽

Tetsuro Fujimoto ◽

...

Keyword(s):

Monoclonal Antibody ◽

Cell Lines ◽

Immunohistochemical Analysis ◽

Specific Protein ◽

Platelet Membrane ◽

Rt Pcr ◽

Chain Reaction ◽

Flow Cytometric ◽

Polymerase Chain ◽

Megakaryocytic Cell

SummaryGlycoprotein V (GPV) is a platelet membrane protein with a molecular weight of 82 kD, and one of the leucine rich glycoproteins (LRG). By reverse transcription-polymerase chain reaction (RT-PCR), GPV cDNA was amplified from mRNA of platelets and megakaryocytic cell lines. However, since there are few reports indicating whether GPV protein is expressed in megakaryocytes as a lineage and maturation specific protein, we studied the GPV expression at the protein level by using a novel monoclonal antibody (1D9) recognizing GPV. Flow cytometric and immunohistochemical analysis indicated that GPV was detected on the surface and in the cytoplasm of only the megakaryocytes in bone marrow aspirates. In a megakaryocytic cell line UT-7, GPV antigen increased after treatment with phorbol-12-myri-state-13-acetate (PMA). These data indicate that only megakaryocytes specifically express the GPV protein among hematopoietic cells and that the expression of GPV increases with differentiation of the megakaryocyte as GPIb-IX complex.

Download Full-text

Expression of type one cannabinoid receptor in different subpopulation of kisspeptin neurons and kisspeptin afferents to GnRH neurons in female mice

Brain Structure and Function ◽

10.1007/s00429-021-02339-z ◽

2021 ◽

Author(s):

Tamás Wilheim ◽

Krisztina Nagy ◽

Mahendravarman Mohanraj ◽

Kamil Ziarniak ◽

Masahiko Watanabe ◽

...

Keyword(s):

Cannabinoid Receptor ◽

Third Ventricle ◽

Glutamate Transporter ◽

Neuroendocrine Cells ◽

Receptor Protein ◽

Specific Expression ◽

Female Mice ◽

Gnrh Neurons ◽

Periventricular Area ◽

Cycle Dependent

AbstractThe endocannabinoids have been shown to target the afferents of hypothalamic neurons via cannabinoid 1 receptor (CB1) and thereby to influence their excitability at various physiological and/or pathological processes. Kisspeptin (KP) neurons form afferents of multiple neuroendocrine cells and influence their activity via signaling through a variation of co-expressed classical neurotransmitters and neuropeptides. The differential potency of endocannabinoids to influence the release of classical transmitters or neuropeptides, and the ovarian cycle-dependent functioning of the endocannabinoid signaling in the gonadotropin-releasing hormone (GnRH) neurons initiated us to study whether (a) the different subpopulations of KP neurons express CB1 mRNAs, (b) the expression is influenced by estrogen, and (c) CB1-immunoreactivity is present in the KP afferents to GnRH neurons. The aim of the study was to investigate the site- and cell-specific expression of CB1 in female mice using multiple labeling in situ hybridization and immunofluorescent histochemical techniques. The results support that CB1 mRNAs are expressed by both the GABAergic and glutamatergic subpopulations of KP neurons, the receptor protein is detectable in two-thirds of the KP afferents to GnRH neurons, and the expression of CB1 mRNA shows an estrogen-dependency. The applied estrogen-treatment, known to induce proestrus, reduced the level of CB1 transcripts in the rostral periventricular area of the third ventricle and arcuate nucleus, and differently influenced its co-localization with vesicular GABA transporter or vesicular glutamate transporter-2 in KP neurons. This indicates a gonadal cycle-dependent role of endocannabinoid signaling in the neuronal circuits involving KP neurons.

Download Full-text

Expression of beta-nerve growth factor and its receptor in rat seminiferous epithelium: specific function at the onset of meiosis.

The Journal of Cell Biology ◽

10.1083/jcb.117.3.629 ◽

1992 ◽

Vol 117 (3) ◽

pp. 629-641 ◽

Cited By ~ 63

Author(s):

M Parvinen ◽

M Pelto-Huikko ◽

O Söder ◽

R Schultz ◽

A Kaipia ◽

...

Keyword(s):

Plasma Membrane ◽

Nerve Growth Factor ◽

Growth Factor ◽

Sertoli Cells ◽

Seminiferous Epithelium ◽

Receptor Protein ◽

Tyrosine Kinase Receptor ◽

Specific Expression ◽

Nerve Growth ◽

Ngf Receptor

beta-Nerve growth factor (NGF) is expressed in spermatogenic cells and has testosterone-downregulated low-affinity receptors on Sertoli cells suggesting a paracrine role in the regulation of spermatogenesis. An analysis of the stage-specific expression of NGF and its low affinity receptor during the cycle of the seminiferous epithelium in the rat revealed NGF mRNA and protein at all stages of the cycle. Tyrosine kinase receptor (trk) mRNA encoding an essential component of the high-affinity NGF receptor was also present at all stages. In contrast, expression of low affinity NGF receptor mRNA was only found in stages VIIcd and VIII of the cycle, the sites of onset of meiosis. The low-affinity NGF receptor protein was present in the plasma membrane of the apical Sertoli cell processes as well as in the basal plasma membrane of these cells at stages VIIcd to XI. NGF was shown to stimulate in vitro DNA synthesis of seminiferous tubule segments with preleptotene spermatocytes at the onset of meiosis while other segments remained nonresponsive. We conclude that NGF is a meiotic growth factor that acts through Sertoli cells.

Download Full-text

Stage-specific Localization and Expression of c-kit in the Adult Human Testis

Journal of Histochemistry & Cytochemistry ◽

10.1369/jhc.2009.953737 ◽

2009 ◽

Vol 57 (9) ◽

pp. 861-869 ◽

Cited By ~ 41

Author(s):

Sreepoorna K. Unni ◽

Deepak N. Modi ◽

Shilpa G. Pathak ◽

Jayesh V. Dhabalia ◽

Deepa Bhartiya

Keyword(s):

Current Knowledge ◽

Protein Localization ◽

Immunohistochemical Analysis ◽

Human Testis ◽

Specific Expression ◽

Adult Human ◽

Kit Receptor ◽

Specific Localization ◽

Human Spermatogenesis ◽

And Function

The c-kit receptor (KIT) and its ligand, stem cell factor (SCF), represent one of the key regulators of testicular formation, development, and function and have been extensively studied in various animal models. The present study was undertaken to characterize the pattern of localization and expression of c-kit in normal adult human testis. Immunohistochemical analysis showed that KIT is expressed in the cytoplasm of spermatogonia, acrosomal granules of spermatids, and Leydig cells. Interestingly, a rather heterogenous pattern of expression of the protein along the basement membrane was observed. Intense protein localization in spermatogonia was detected in stages I–III, whereas low expression was observed in stages IV–VI of the seminiferous epithelium, indicating that the expression of the molecule was stage specific. In situ hybridization studies revealed that the transcripts of the gene were also localized in a similar non-uniform pattern. To the best of our knowledge, such a stage-specific expression of KIT has not been reported previously in the human testis. The results of the present study may expand current knowledge about the c-kit/SCF system in human spermatogenesis.

Download Full-text

Differential mucin gene expression in human pancreatic and colon cancer cells

Biochemical Journal ◽

10.1042/bj2760599 ◽

1991 ◽

Vol 276 (3) ◽

pp. 599-605 ◽

Cited By ~ 42

Author(s):

S Yonezawa ◽

J C Byrd ◽

R Dahiya ◽

J J L Ho ◽

J R Gum ◽

...

Keyword(s):

Pancreatic Cancer ◽

Colon Cancer ◽

Cancer Cells ◽

Cell Lines ◽

Cancer Cell ◽

Cancer Cell Lines ◽

Human Pancreatic Cancer ◽

Northern Blots ◽

Western Blots ◽

Pancreatic Cancer Cells

The purpose of this study was to determine the quantity and nature of the mucins synthesized and secreted by four different pancreatic cancer cell lines. Well- to moderately-differentiated SW1990 and CAPAN-2 human pancreatic cancer cells were found to produce more high-Mr glycoprotein (HMG) than less-differentiated MIA PaCa-2 and PANC-1 cells. Most of the labelled HMG was secreted within 24 h. The results of chemical and enzymic degradation, ion-exchange chromatography and density-gradient centrifugation indicated that the HMG in SW1990 and CAPAN-2 cells has the properties expected for mucins, whereas much of the HMG in MIA PaCa-2 and PANC-1 cells may not be mucin, but proteoglycan. These results are consistent with immunoblots and Northern blots showing the presence of apomucin and apomucin mRNA in SW1990 and CAPAN-2 cells, but not in MIA PaCa-2 and PANC-1 cells. The Western blots and Northern blots also show that SW1990 and CAPAN-2 cells, like breast cancer cells, have the mammary-type apomucin and mRNA coded by the MUC1 gene, but lack the intestinal type apomucin and mRNA coded by the MUC2 gene. In contrast, the colon cancer cell lines tested in culture express apomucin and mRNA coded by MUC2 but not by MUC1.

Download Full-text

Abstract 269: Miofibroblast Tgf-beta Signaling in a Proteotoxic Mouse Model

Circulation Research ◽

10.1161/res.119.suppl_1.269 ◽

2016 ◽

Vol 119 (suppl_1) ◽

Author(s):

Bidur Bhandary ◽

Qinghang Meng ◽

Hanna Osinska ◽

Kritton Shay-Winkler ◽

James Gulick ◽

...

Keyword(s):

Heart Disease ◽

Mouse Model ◽

Cardiac Function ◽

Transforming Growth Factor Beta ◽

Cardiac Fibrosis ◽

Transforming Growth Factor ◽

Prolonged Survival ◽

Tgfβ Signaling ◽

Specific Expression ◽

Western Blots

Introduction: Transforming Growth Factor Beta (TGFβ) is an important cytokine in mediating the fibrogenic response and, in particular, cardiac fibrosis. Extensive fibrosis accompanies the cardiac remodeling that occurs during development of the protein conformation-based disease caused by cardiomyocyte-specific expression of a mutant, small, heat shock-like protein and chaperone, aB crystallin (CryABR120G). During the onset of fibrosis, fibroblasts are activated to the so-called “myofibroblast” state and TGFβ binding is thought to mediate an essential signaling pathway underlying this process. Our central hypothesis is that TGFβ signaling processes that result in significant cardiac fibrosis in a mouse model of proteotoxic heart disease are mediated by cardiac fibroblasts, rather than cardiomyocytes. Here, we have partially ablated TGFβ signaling only in cardiac myofibroblasts to observe if cardiac fibrosis is reduced. Aims and Methods: The objective of this study was to understand the contributions of fibroblast-derived TGFβ signaling to the development of cardiac fibrosis in a proteotoxic mouse model that results in significant cardiac fibrosis. To test the hypothesis we partially deleted the myofibroblast specific canonical and non-canonical signaling by crossing CryAB R120G mice with Tgfbr1 or Tgfbr2 floxed mice. The double transgene containing mice were further crossed with activated myofibroblast specific Cre mice in which Cre expression was driven off the periostin promoter. Echocardiography, Masson’s Trichome staining, PCR arrays, IHC and western blots were performed to characterize the fibrotic progression in CryAB R120G transgenic mice. Results: We observed that myofibroblast-targeted partial knockdown of Tgf βr1 signaling prolonged survival, modestly reducing fibrosis and improving cardiac function . Similarly, Tgf βr2 partial knockdown prolonged survival, modestly reducing fibrosis without improving cardiac function during fibrosis development in CryAB R120G mice. Conclusion: These findings suggest that, in a model of proteotoxic heart disease, myofibroblast based TGFβ signaling in the heart may contribute to cardiac hypertrophy/dysfunction but cannot account entirely for the fibrotic response.

Download Full-text

Abstract 235: Sarcolemmal Membrane Associated Protein Isoform 1: a Unique Regulator of Glucose Uptake and Metabolism in the Myocardium

Circulation Research ◽

10.1161/res.117.suppl_1.235 ◽

2015 ◽

Vol 117 (suppl_1) ◽

Author(s):

Aaraf Dewan ◽

Maysoon Salih ◽

Christopher Triggle ◽

Hong Ding ◽

Balwant Tuana

Keyword(s):

Glucose Uptake ◽

Glucose Transporter ◽

Glucose Transporter 4 ◽

Specific Expression ◽

Western Blots ◽

Sarcolemmal Membrane ◽

Neonatal Cardiomyocytes ◽

Early Endosomes ◽

Protein Isoform ◽

Uptake And Metabolism

As one of the leading causes of heart disease, diabetes is a problem which needs a solution. Regulation of glucose uptake and metabolism within skeletal and cardiac muscle has proven capable of altering systemic glucose levels and impacting metabolism to potentially improve patient outcomes. Unfortunately, to date, very few muscle specific metabolic regulators are known which can allow us to achieve blood glucose uptake and metabolism. Sarcolemmal Membrane Associated Protein Isoform 1 (SLMAP1) is a novel protein expressed predominantly within muscle tissue. It has been linked to diabetes through animal models, although its role in metabolism remains to be defined. Here we describe a novel role for SLMAP1 in glucose metabolism within the myocardium. We engineered a transgenic (TG) mouse with cardiac specific expression of SLMAP1. Using neonatal cardiomyocytes (NCMs) collected from these mice we performed glucose uptake assays with 2-deoxy-glucose (2DG), measured glycolytic rate using an Extracellular Flux XF24e Bioanalyzer, and analyzed glucose transporter 4 (GLUT4) trafficking using Western Blots, qPCR, and immunofluorescence imaging. NCMs extracted from TG hearts expressing SLMAP1 displayed increased levels of 2DG uptake (93% ± 25%, n=5, P<0.01), basal glycolysis (glycolysis (92 ± 40%, n=5, P<0.05), and maximal glycolysis (75 ± 31%, n=5, P<0.05) compared with wildtype littermates. Confocal microscopy revealed both increased localization of glucose transporter 4 (GLUT4) at the cell surface as well as an expansion of GLUT4 early endosomes in TG NCMs. The data here indicates SLMAP1 as a novel regulator of glucose uptake and metabolism in the myocardium. The targeted expression of SLMAP1 in a muscle specific manner may enhance systemic glycemic control and serve to limit cardiovascular disease in metabolic disorders such as diabetes.

Download Full-text

Developmental stage- and spermatogenic cycle-specific expression of transcription factor GATA-1 in mouse Sertoli cells

Development ◽

10.1242/dev.120.7.1759 ◽

1994 ◽

Vol 120 (7) ◽

pp. 1759-1766 ◽

Cited By ~ 6

Author(s):

K. Yomogida ◽

H. Ohtani ◽

H. Harigae ◽

E. Ito ◽

Y. Nishimune ◽

...

Keyword(s):

Transcription Factor ◽

Developmental Stage ◽

Sertoli Cell ◽

Germ Cells ◽

Sertoli Cells ◽

Transcriptional Activation ◽

Germ Line ◽

Mouse Testis ◽

Seminiferous Tubules ◽

Specific Expression

GATA-1 is an essential factor for the transcriptional activation of erythroid-specific genes, and is also abundantly expressed in a discrete subset of cells bordering the seminiferous epithelium in tubules of the murine testis. In examining normal and germ-line defective mutant mice, we show here that GATA-1 is expressed only in the Sertoli cell lineage in mouse testis. GATA-1 expression in Sertoli cells is induced concomitantly with the first wave of spermatogenesis, and GATA-1-positive cells are uniformly distributed among all tubules during prepubertal testis development. However, the number of GATA-1-positive cells declines thereafter and were found only in the peripheral zone of seminiferous tubules in stages VII, VIII and IX of spermatogenesis in the adult mouse testis. In contrast, virtually every Sertoli cell in mutant W/Wv, jsd/jsd or cryptorchid mice (all of which lack significant numbers of germ cells) expresses GATA-1, thus showing that the expression of this transcription factor is negatively controlled by the maturing germ cells. These observations suggest that transcription factor GATA-1 is a developmental stage- and spermatogenic cycle-specific regulator of gene expression in Sertoli cells.

Download Full-text

Cell proliferation contributes to PNEC hyperplasia after acute airway injury

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.1997.272.3.l486 ◽

1997 ◽

Vol 272 (3) ◽

pp. L486-L493 ◽

Cited By ~ 9

Author(s):

T. P. Stevens ◽

J. T. McBride ◽

J. L. Peake ◽

K. E. Pinkerton ◽

B. R. Stripp

Keyword(s):

Cell Proliferation ◽

Epithelial Cells ◽

Metabolic Activation ◽

Immunohistochemical Analysis ◽

Airway Epithelial Cells ◽

Neuroendocrine Cells ◽

Clara Cells ◽

Proliferating Cells ◽

Airway Injury ◽

Chronic Lung Diseases

Pulmonary neuroendocrine cells (PNECs) are airway epithelial cells that are capable of secreting a variety of neuropeptides. PNECs are scattered throughout the bronchial tree either as individual cells or clusters of cells termed neuroepithelial bodies (NEBs). PNECs and their secretory peptides have been considered to play a role in fetal lung development. Although the normal physiological function of PNECs and neuropeptides in normal adult lungs and in repair from lung injury is not known, PNEC hyperplasia has been associated with chronic lung diseases, such as bronchopulmonary dysplasia, and with chronic exposures, such as hypoxia, tobacco smoke, nitrosamines, and ozone. To evaluate changes in PNEC number and distribution after acute airway injury, FVB/n mice were treated with either naphthalene or vehicle. Naphthalene is an aromatic hydrocarbon that, at the dose used in this study, selectively destroys nonciliated bronchial epithelial cells (Clara cells) through cytochrome P-450-mediated metabolic activation into cytotoxic epoxides. PNECs were identified by immunohistochemical analysis of calcitonin gene-related peptide-like immunoreactivity (CGRP-IR). Proliferating cells were marked with [(3)H]thymidine incorporation. Acute naphthalene toxicity results in PNEC hyperplasia that is detectable after 5 days of recovery. PNEC hyperplasia is characterized by increased numbers of NEBs without significant changes in the number of isolated PNECs and by increased [(3)H]thymidine labeling of CGRP-IR cells. These data show that cell proliferation contributes to PNEC hyperplasia after acute airway injury and suggest that PNECs may be capable of more rapidly increasing their number in response to injury than previously recognized.

Download Full-text