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Testicular Connexin43 (Cx43) connects adjacent Sertoli cells (SC) and SC to germ cells (GC) in the seminiferous epithelium and plays a crucial role in spermatogenesis. However, the distinction whether this results from impaired inter-SC communication or between GC and SC is not possible, so far. Thus, the question arises, whether a GC-specific Cx43 KO has similar effects on spermatogenesis as it is general or SC-specific KO. Using the Cre/loxP recombinase system, two conditional KO mouse lines lacking Cx43 in premeiotic (pGCCx43KO) or meiotic GC (mGCCx43KO) were generated. It was demonstrated by qRT-PCR that Cx43 mRNA was significantly decreased in adult pGCCx43KO mice, while it was also reduced in mGCCx43KO mice, yet not statistically significant. Body and testis weights, testicular histology, tubular diameter, numbers of intratubular cells and Cx43 protein synthesis and localization did not show any significant differences in semi-quantitative Western blot analysis and immunohistochemistry comparing adult male KO and WT mice of both mouse lines. Male KO mice were fertile. These results indicate that Cx43 in spermatogonia/spermatids does not seem to be essential for successful termination of spermatogenesis and fertility as it is known for Cx43 in somatic SC, but SC-GC communication might rather occur via heterotypic GJ channels.
Kristina Rode; Marion Langeheine; Bettina Seeger; Ralph Brehm. Connexin43 in Germ Cells Seems to Be Dispensable for Murine Spermatogenesis. International Journal of Molecular Sciences 2021, 22, 7924 .
AMA StyleKristina Rode, Marion Langeheine, Bettina Seeger, Ralph Brehm. Connexin43 in Germ Cells Seems to Be Dispensable for Murine Spermatogenesis. International Journal of Molecular Sciences. 2021; 22 (15):7924.
Chicago/Turabian StyleKristina Rode; Marion Langeheine; Bettina Seeger; Ralph Brehm. 2021. "Connexin43 in Germ Cells Seems to Be Dispensable for Murine Spermatogenesis." International Journal of Molecular Sciences 22, no. 15: 7924.
The wild boar population in Europe is steadily growing, one of the reasons for this increase probably being the high reproductive potential of this large mammal. Population management is important to stabilise wild boar numbers and a great deal of attention is focusing on the reasons, which might contribute to the high reproductive rates. Understanding the timing of puberty attainment provides information required for proper management practices. Knowledge of the earliest expected time of sexual maturation in male wild boars is limited, research being mostly focused on females. Previous hunting references indicate that sexual maturity in males occurs in the second year after birth. In contrast, male domestic pigs become sexually mature from about seven months of age. Thus, aims of this study were to investigate (1) whether there is a physiological ability for reproduction also in male wild boars of a younger age and (2) whether the body weight of wild boar males has a more important role than age in driving the maturation of the testis. Male wild boar individuals were sampled during hunting drives in the eastern part of Lower Saxony in Germany. Testes with epididymides from 74 males were collected and prepared for histological examination and immunohistochemistry. The reproductive status could be ascertained based on development/occurrence of different germ cell populations using histology and based on the immunohistochemical detection of the anti-Müllerian hormone and androgen receptor. In this study, male wild boars aged nine to ten months already passed puberty and were able to reproduce if they had reached the appropriate body condition of about 29 kg dressed weight. Immunopositivity to the anti-Müllerian hormone in Sertoli cells was evident only in prepubertal animals and decreased with the onset of puberty. No immunoreaction was evident at postpuberty. The androgen receptor was detected in Sertoli cells, peritubular cells and Leydig cells, surprisingly already in Sertoli cells of prepubertal wild boars as well depending on body weight. Moreover, two-thirds of young males aged about ten months were precociously reproductively mature, showing histologically the presence of spermatozoa in testes and epididymides. As piglets are mostly born in spring, also these young male individuals could target the heat of female wild boars in the winter months, resulting in the observed population increase. Therefore, a reduction in wild boar numbers should also focus on piglets of both sexes.
Claudia Maistrelli; Hanna Hüneke; Marion Langeheine; Oliver Keuling; Ursula Siebert; Ralph Brehm. Precocious puberty in male wild boars: a possible explanation for the dramatic population increase in Germany and Europe. PeerJ 2021, 9, e11798 .
AMA StyleClaudia Maistrelli, Hanna Hüneke, Marion Langeheine, Oliver Keuling, Ursula Siebert, Ralph Brehm. Precocious puberty in male wild boars: a possible explanation for the dramatic population increase in Germany and Europe. PeerJ. 2021; 9 ():e11798.
Chicago/Turabian StyleClaudia Maistrelli; Hanna Hüneke; Marion Langeheine; Oliver Keuling; Ursula Siebert; Ralph Brehm. 2021. "Precocious puberty in male wild boars: a possible explanation for the dramatic population increase in Germany and Europe." PeerJ 9, no. : e11798.
Rye could offer diverse benefits in terms of sustainability if it could replace parts of the main cereals, corn and wheat, in broiler diets. A total of 256 broilers, Ross 308, were randomly allocated into 32 pens. From day 14 till day 42, the birds were divided into four feeding groups (eight replicates each). The control group received a conventional finisher diet “control”, whereas in the other groups, a pelleted supplementary feed was offered (SFI to corn and SFII to rye), to which crushed corn (SFI-Corn) or squashed rye (SFII-Rye) was added. The fourth group received a mixture of 50% SFI-Corn and 50% SFII-Rye. The cereal level was increased weekly (5%, 10%, 20%, 30%) at the expense of the supplementary feeds. No significant effects were observed for body weight at d 42 and excreta viscosity between all groups. Overall, foot pad health was excellent. Compared to the control group, birds fed SFI-Corn displayed a significant increase in gizzard relative weight, whereas, in contrast to all other groups, ileal villus height was significantly lower. In conclusion, feeding SFI-Corn or SFII-Rye diets had no negative influences on performance, litter quality and digesta viscosity, whereas SFI-Corn partially affected ileal morphology.
Amr El-Wahab; Jan Lingens; Bussarakam Chuppava; Marwa Ahmed; Ahmed Osman; Marion Langeheine; Ralph Brehm; Venja Taube; Richard Grone; Andreas Von Felde; Josef Kamphues; Christian Visscher. Impact of Rye Inclusion in Diets for Broilers on Performance, Litter Quality, Foot Pad Health, Digesta Viscosity, Organ Traits and Intestinal Morphology. Sustainability 2020, 12, 7753 .
AMA StyleAmr El-Wahab, Jan Lingens, Bussarakam Chuppava, Marwa Ahmed, Ahmed Osman, Marion Langeheine, Ralph Brehm, Venja Taube, Richard Grone, Andreas Von Felde, Josef Kamphues, Christian Visscher. Impact of Rye Inclusion in Diets for Broilers on Performance, Litter Quality, Foot Pad Health, Digesta Viscosity, Organ Traits and Intestinal Morphology. Sustainability. 2020; 12 (18):7753.
Chicago/Turabian StyleAmr El-Wahab; Jan Lingens; Bussarakam Chuppava; Marwa Ahmed; Ahmed Osman; Marion Langeheine; Ralph Brehm; Venja Taube; Richard Grone; Andreas Von Felde; Josef Kamphues; Christian Visscher. 2020. "Impact of Rye Inclusion in Diets for Broilers on Performance, Litter Quality, Foot Pad Health, Digesta Viscosity, Organ Traits and Intestinal Morphology." Sustainability 12, no. 18: 7753.
The aim of this study was to test whether a single testicular needle biopsy could provide histological results comparable to en bloc resection histology and whether one biopsy was sufficient to reflect the histology of an entire pair of testicles. Two methods of sample collection were tested on 32 bull calves aged five to eight months to compare histological parameters of needle biopsy with those of en bloc resection samples. One testicular needle biopsy of the right and three en bloc samples of both testicles were collected and compared for the number of tubular cross sections, tubules with elongated spermatids (ES), outer/inner diameter of tubules, thickness of tubular wall, and number of Sertoli cells (SC). Additionally, animal data were considered. No significant differences were found between the left and right testis or among the individual locations of en bloc samples. However, histologically significant differences (Bonferroni-adjusted significance level: p < 0.05/6 = 0.0083) were found between the needle biopsy and en bloc resection regarding the tubular cross sections per visual field (p < 0.05), the outer (p = 0.01) and inner diameter and the thickness of the tubular wall (both p < 0.01). In the SOX9 immunohistochemical staining, no significant differences (p > 0.05) could be observed for SC numbers between needle biopsy and en bloc samples. In conclusion, results of testicular needle biopsy do not have the same validity as the en bloc resection histology. Furthermore, one biopsy is insufficient to reflect the histology of the entire testicular pair.
Maike Rohländer; Henning Otzen; Kristina Rode; Klaus Jung; Marion Schmicke; Teresa Harborth; Marion Langeheine; Ralph Brehm; Árpád Csaba Bajcsy. Histological Comparison of Testicular Needle Biopsy and En Bloc Samples in Abattoir Calves. Animals 2020, 10, 918 .
AMA StyleMaike Rohländer, Henning Otzen, Kristina Rode, Klaus Jung, Marion Schmicke, Teresa Harborth, Marion Langeheine, Ralph Brehm, Árpád Csaba Bajcsy. Histological Comparison of Testicular Needle Biopsy and En Bloc Samples in Abattoir Calves. Animals. 2020; 10 (5):918.
Chicago/Turabian StyleMaike Rohländer; Henning Otzen; Kristina Rode; Klaus Jung; Marion Schmicke; Teresa Harborth; Marion Langeheine; Ralph Brehm; Árpád Csaba Bajcsy. 2020. "Histological Comparison of Testicular Needle Biopsy and En Bloc Samples in Abattoir Calves." Animals 10, no. 5: 918.
The Sertoli cell (SC) specific connexin43 (Cx43) knockout (SCCx43KO) mouse line is ideal to gain insight into the mechanistic gap junction formation in SC and the seminiferous epithelium. A method for developing primary SC cultures from these mice was established, validated and successfully characterized via polymerase chain reaction, immunohistochemistry, immunofluorescence (IF), and Western blots (WB). It was evident that both knockout (KO) and wild-type (WT) primary cell cultures were similar in morphology. These highly pure SC cultures were subjected to cell proliferation assays indicating no notable proliferation in cultures of both genotypes. Measurements of cell monolayer integrity indicated significant increases in transepithelial electrical resistance and consequently in tight junction expression of the KO cultures. Using semi-quantitative WB and IF, tight junction protein claudin-11 was analyzed. These results support a role for Cx43 in regulating blood-testis barrier (BTB) function, composition, and dynamics in vitro. Thus, the SC deficient Cx43 cell cultures may provide a valuable in vitro tool for a better understanding of the mechanistic role of Cx43 in spermatogenesis and BTB assembly.
Jonathan Gerber; Kristina Rode; Nina Hambruch; Marion Langeheine; Nadine Schnepel; Ralph Brehm. Establishment and functional characterization of a murine primary Sertoli cell line deficient of connexin43. Cell and Tissue Research 2020, 381, 309 -326.
AMA StyleJonathan Gerber, Kristina Rode, Nina Hambruch, Marion Langeheine, Nadine Schnepel, Ralph Brehm. Establishment and functional characterization of a murine primary Sertoli cell line deficient of connexin43. Cell and Tissue Research. 2020; 381 (2):309-326.
Chicago/Turabian StyleJonathan Gerber; Kristina Rode; Nina Hambruch; Marion Langeheine; Nadine Schnepel; Ralph Brehm. 2020. "Establishment and functional characterization of a murine primary Sertoli cell line deficient of connexin43." Cell and Tissue Research 381, no. 2: 309-326.
Male factor infertility is a problem in today’s society but many underlying causes are still unknown. The generation of a conditional Sertoli cell (SC)-specific connexin 43 (Cx43) knockout mouse line (SCCx43KO) has provided a translational model. Expression of the gap junction protein Cx43 between adjacent SCs as well as between SCs and germ cells (GCs) is known to be essential for the initiation and maintenance of spermatogenesis in different species and men. Adult SCCx43KO males show altered spermatogenesis and are infertile. Thus, the present study aims to identify molecular mechanisms leading to testicular alterations in prepubertal SCCx43KO mice. Transcriptome analysis of 8-, 10- and 12-day-old mice was performed by next-generation sequencing (NGS). Additionally, candidate genes were examined by qRT-PCR and immunohistochemistry. NGS revealed many significantly differentially expressed genes in the SCCx43KO mice. For example, GC-specific genes were mostly downregulated and found to be involved in meiosis and spermatogonial differentiation (e.g., Dmrtb1, Sohlh1). In contrast, SC-specific genes implicated in SC maturation and proliferation were mostly upregulated (e.g., Amh, Fshr). In conclusion, Cx43 in SCs appears to be required for normal progression of the first wave of spermatogenesis, especially for the mitosis-meiosis switch, and also for the regulation of prepubertal SC maturation.
Erika Hilbold; Ottmar Distl; Martina Hoedemaker; Sandra Wilkening; Rüdiger Behr; Aleksandar Rajkovic; Marion Langeheine; Kristina Rode; Klaus Jung; Julia Metzger; Ralph H. J. Brehm. Loss of Cx43 in Murine Sertoli Cells Leads to Altered Prepubertal Sertoli Cell Maturation and Impairment of the Mitosis-Meiosis Switch. Cells 2020, 9, 676 .
AMA StyleErika Hilbold, Ottmar Distl, Martina Hoedemaker, Sandra Wilkening, Rüdiger Behr, Aleksandar Rajkovic, Marion Langeheine, Kristina Rode, Klaus Jung, Julia Metzger, Ralph H. J. Brehm. Loss of Cx43 in Murine Sertoli Cells Leads to Altered Prepubertal Sertoli Cell Maturation and Impairment of the Mitosis-Meiosis Switch. Cells. 2020; 9 (3):676.
Chicago/Turabian StyleErika Hilbold; Ottmar Distl; Martina Hoedemaker; Sandra Wilkening; Rüdiger Behr; Aleksandar Rajkovic; Marion Langeheine; Kristina Rode; Klaus Jung; Julia Metzger; Ralph H. J. Brehm. 2020. "Loss of Cx43 in Murine Sertoli Cells Leads to Altered Prepubertal Sertoli Cell Maturation and Impairment of the Mitosis-Meiosis Switch." Cells 9, no. 3: 676.
The blood-testis barrier (BTB) consists of different cell-to-cell connections, including tight junction proteins like claudin-11 (CLDN11). For dogs, only limited data is published dealing with these proteins in general. Therefore, their physiological relevance, their postnatal expression, and their distribution pattern in pathological conditions, e.g. in altered spermatogenesis and testicular neoplasia were assessed. Canine testes from routine castrations, and those sent in for diagnostic purposes were investigated. Based on morphological evaluation, the dogs and testes were divided into groups: (1) dogs with normal spermatogenesis, (2) four months old prepubertal dogs, (3) intratubular seminoma, (4) diffuse seminoma, (5) Sertoli cell tumours (SCT), (6) Leydig cell tumours (LCT), and (7) dogs with impaired spermatogenesis (e.g. mixed atrophy). In order to examine possible alterations of the BTB components, immunohistochemistry (IHC) and immunofluorescence using a commercial antibody against CLDN11 was performed. Sertoli cell (SC) nuclei (SOX9) and peritubular myoid cells (smooth-muscle-actin, SMA) were also assessed using IHC. Additionally, semi-quantitative Western-blot (WB) and RT-PCR analyses of CLDN11 were conducted. In tubules with normal spermatogenesis, IHC of CLDN11 revealed a basolateral staining at BTB localisation. In prepubertal cords, CLDN11 was diffusely expressed along the cytoplasmic extensions of SCs supposing that the BTB was neither built up nor functional, yet. A shift from weakly expressed CLDN11 between/in residual SCs in intratubular seminoma to only small CLDN11 immunopositive stained spots in the cytoplasm of remaining SOX9-positive SCs in diffuse seminoma was detectable. Reduction or even loss of CLDN11 expression in diffuse seminoma was confirmed using RT-PCR and WB analyses, thus indicating that in seminoma, CLDN11 was downregulated at transcriptional level and completely lost its sealing function. Basal SCs in SCT still showed a CLDN11/SOX9 co-localisation, suggesting that luminal neoplastic SCs undergo de-differentiation during tumour progression. In LCT, no CLDN11 was detectable. Dogs with mixed atrophy showed an upregulation of CLDN11 in tubules with spermatogenic arrest on mRNA and protein level, leading to the conclusion that within these tubules regulatory mechanisms lost their equilibrium. For the first time, the spatial expression of CLDN11 in prepubertal canine testis, impaired spermatogenesis, intratubular seminoma and its absence in diffuse seminoma and LCT was shown. Since altered CLDN11 levels could be part of adaptive mechanisms to modify BTB integrity, further functional investigations to characterize the canine BTB need to be conducted.
Carolin Pörtner; Kristina Rode; Julia Hollenbach; Heike Thiemeyer; Andreas Beineke; Anne-Rose Günzel-Apel; Ralph Brehm. Expression of claudin-11 in canine prepubertal testes, and in canine adult testes showing normal spermatogenesis, impaired spermatogenesis, or testicular neoplasia. Theriogenology 2020, 148, 122 -131.
AMA StyleCarolin Pörtner, Kristina Rode, Julia Hollenbach, Heike Thiemeyer, Andreas Beineke, Anne-Rose Günzel-Apel, Ralph Brehm. Expression of claudin-11 in canine prepubertal testes, and in canine adult testes showing normal spermatogenesis, impaired spermatogenesis, or testicular neoplasia. Theriogenology. 2020; 148 ():122-131.
Chicago/Turabian StyleCarolin Pörtner; Kristina Rode; Julia Hollenbach; Heike Thiemeyer; Andreas Beineke; Anne-Rose Günzel-Apel; Ralph Brehm. 2020. "Expression of claudin-11 in canine prepubertal testes, and in canine adult testes showing normal spermatogenesis, impaired spermatogenesis, or testicular neoplasia." Theriogenology 148, no. : 122-131.
Sustainably produced poultry meat with consideration of animal health poses a challenge for broiler production. Low protein diets with high amounts of synthetic amino acids (AAs) like methionine (Met) are the consequence. In a five-week feeding trial, 360 broilers (Ross 308) assigned to four feeding groups were offered protein-reduced complete diets (starter: 20% crude protein (CP); grower: 18.5% CP; finisher: 17.5% CP), supplemented with essential AAs. The “MHA” group received DL-2-hydroxy-4-(methylthio) butanoic acid (DL-HMTBA; trade name: MHA®), groups “L” and “DL” the respective Met source in equivalent concentrations each exceeding the nutritional recommendations. “R-MHA” (“R” for “reduced”) received the minimum required level (using MHA as Met source). Performance exceeded performance goals without differences between the groups. The average feed conversion ratio (FCR) amounted to 1.35. The carcass/body weight ratio of R-MHA was significantly lower (0.782) compared to DL (0.808) and L (0.809). Breast meat of R-MHA contained significantly more fat (144 g/kg dry matter (DM)) compared to L (104 g/kg DM) and significantly lower CP content (R-MHA: 838 g/kg DM; L: 875 g/kg DM). The results indicated possible improvement in slaughter yield by protein-reduced diets supplemented with L-Met, thus recommending further research focusing on the Met influence.
Cristina Ullrich; Marion Langeheine; Ralph Brehm; Venja Taube; Mercedes Rosillo Galera; Karl Rohn; Johanna Popp; Christian Visscher. Influence of Different Methionine Sources on Performance and Slaughter Characteristics of Broilers. Animals 2019, 9, 984 .
AMA StyleCristina Ullrich, Marion Langeheine, Ralph Brehm, Venja Taube, Mercedes Rosillo Galera, Karl Rohn, Johanna Popp, Christian Visscher. Influence of Different Methionine Sources on Performance and Slaughter Characteristics of Broilers. Animals. 2019; 9 (11):984.
Chicago/Turabian StyleCristina Ullrich; Marion Langeheine; Ralph Brehm; Venja Taube; Mercedes Rosillo Galera; Karl Rohn; Johanna Popp; Christian Visscher. 2019. "Influence of Different Methionine Sources on Performance and Slaughter Characteristics of Broilers." Animals 9, no. 11: 984.
Respiratory infections caused by mycoplasma species in ruminants lead to considerable economic losses. Two important ruminant pathogens are Mycoplasma mycoides subsp. Mycoides (Mmm), the aetiological agent of contagious bovine pleuropneumonia and Mycoplasma mycoides subsp. capri (Mmc), which causes pneumonia, mastitis, arthritis, keratitis, and septicemia in goats. We established precision cut lung slices (PCLS) infection model for Mmm and Mmc to study host-pathogen interactions. We monitored infection over time using immunohistological analysis and electron microscopy. Moreover, infection burden was monitored by plating and quantitative real-time PCR. Results were compared with lungs from experimentally infected goats and cattle. Lungs from healthy goats and cattle were also included as controls. PCLS remained viable for up to two weeks. Both subspecies adhered to ciliated cells. However, the titer of Mmm in caprine PCLS decreased over time, indicating species specificity of Mmm. Mmc showed higher tropism to sub-bronchiolar tissue in caprine PCLS, which increased in a time-dependent manner. Moreover, Mmc was abundantly observed on pulmonary endothelial cells, indicating partially, how it causes systemic disease. Tissue destruction upon prolonged infection of slices was comparable to the in vivo samples. Therefore, PCLS represents a novel ex vivo model to study host-pathogen interaction in livestock mycoplasma.
Yenehiwot Berhanu Weldearegay; Sandy Müller; Jana Hänske; Anja Schulze; Aline Kostka; Nancy Rüger; Marion Hewicker-Trautwein; Ralph Brehm; Peter Valentin-Weigand; Robert Kammerer; Joerg Jores; Jochen Meens. Host-Pathogen Interactions of Mycoplasma mycoides in Caprine and Bovine Precision-Cut Lung Slices (PCLS) Models. Pathogens 2019, 8, 82 .
AMA StyleYenehiwot Berhanu Weldearegay, Sandy Müller, Jana Hänske, Anja Schulze, Aline Kostka, Nancy Rüger, Marion Hewicker-Trautwein, Ralph Brehm, Peter Valentin-Weigand, Robert Kammerer, Joerg Jores, Jochen Meens. Host-Pathogen Interactions of Mycoplasma mycoides in Caprine and Bovine Precision-Cut Lung Slices (PCLS) Models. Pathogens. 2019; 8 (2):82.
Chicago/Turabian StyleYenehiwot Berhanu Weldearegay; Sandy Müller; Jana Hänske; Anja Schulze; Aline Kostka; Nancy Rüger; Marion Hewicker-Trautwein; Ralph Brehm; Peter Valentin-Weigand; Robert Kammerer; Joerg Jores; Jochen Meens. 2019. "Host-Pathogen Interactions of Mycoplasma mycoides in Caprine and Bovine Precision-Cut Lung Slices (PCLS) Models." Pathogens 8, no. 2: 82.
The transcription factor DMRTB1 plays a pivotal role in coordinating the transition between mitosis and meiosis in murine germ cells. No reliable data are available for human testis. The present study aims to examine the testicular expression pattern of DMRTB1 in men showing normal and impaired spermatogenesis. Immunohistochemistry was performed using 54 human testicular biopsy specimens and a commercial rabbit polyclonal anti-DMRTB1 primary antibody. RT-PCR complemented immunohistochemistry. To further characterize immunopositive cells and possible co-localization, the proliferation marker Ki-67, the tumor marker PLAP, and an anti-DMRT1 antibody were used. In men with normal spermatogenesis, a strong immunoreactivity was detectable in a subset of spermatogonia (38.34 ± 2.14%). Some spermatocytes showed a weak immunostaining. Adjacent Sertoli cells were immunonegative. Compared with a hematoxylin and eosin overview staining, these immunopositive cells were almost exclusively identified as Apale and B spermatogonia and primary spermatocytes in (pre-)leptotene, zygotene, and pachytene stages. In patients with spermatogenic arrest at spermatogonial level, an altered staining pattern was found. No immunoreactivity was detected in Sertoli cells in Sertoli cell-only syndrome. In germ cell neoplasia in situ (GCNIS) tubules, except for a few (0.4 ± 0.03%), pre-invasive tumor cells were immunonegative. Seminoma cells showed no immunostaining. According to previous findings in mice, it seems reasonable that DMRTB1 is expressed in these normal germ cell populations. Moreover, altered staining pattern in spermatogenic arrest at spermatogonial stage suggests a correlation with mitosis and transformation into B spermatogonia. The absence of DMRTB1 in GCNIS cells and tumor cells might be associated with uncontrolled neoplastic cell proliferation and progression into invasive germ cell tumors. Further research is required to elucidate, for example, the role of DMRTB1 in the malignant transformation of human germ cells. Our data indicate a relevant role for DMRTB1 regarding the entry of spermatogonia into meiosis in men.
E. Hilbold; M. Bergmann; D. Fietz; S. Kliesch; W. Weidner; M. Langeheine; K. Rode; R. Brehm. Immunolocalization of DMRTB1 in human testis with normal and impaired spermatogenesis. Andrology 2019, 7, 428 -440.
AMA StyleE. Hilbold, M. Bergmann, D. Fietz, S. Kliesch, W. Weidner, M. Langeheine, K. Rode, R. Brehm. Immunolocalization of DMRTB1 in human testis with normal and impaired spermatogenesis. Andrology. 2019; 7 (4):428-440.
Chicago/Turabian StyleE. Hilbold; M. Bergmann; D. Fietz; S. Kliesch; W. Weidner; M. Langeheine; K. Rode; R. Brehm. 2019. "Immunolocalization of DMRTB1 in human testis with normal and impaired spermatogenesis." Andrology 7, no. 4: 428-440.
Knowledge about reproductive parameters in male harbour porpoises such as testicular histology and germ cell maturation as well as seasonal changes in spermatogenesis is scarce. Thus, the aim of the present study was to report changes in the histological appearance of the testicular morphology of neonatal and juvenile harbour porpoises during maturation, to identify stages of spermatogenesis in adult males and to detect seasonal modifications. The identification of these stages can be used to assess the developmental profile of gene expression during spermatogenesis and to identify defects in spermatogenesis arising in pathological conditions. Testes of adult male harbour porpoises from the North and Baltic Sea that became stranded or by-caught in the years 1998–2016 were histologically examined using Haematoxylin and Eosin – staining. The Periodic Acid Schiff (PAS) staining was used for spermatogenic staging and the evaluation of the development of the acrosomic cap. For the identification of changes in testes morphology and morphometry during the course of the year, histological characteristics like germ cell associations and diameter of the convoluted seminiferous tubules were noted for each month. The analysis showed that in adult males more than one stage of spermatogenesis could be found per cross section of the convoluted seminiferous tubules similar to findings in men and some ape species. This rare phenomenon is called multi-stage-arrangement. In sexually active males from the peak breeding season (June and July) eight stages of spermatogenesis were identified and all stages occurred simultaneously, while during the low breeding season (August to May) only residual spermatogenesis or constituent germ cell populations were found. Missing germ cell generations were recorded in specimens from July to September. Our investigations provide a detailed staging of spermatogenesis and give new insight into the reproductive biology of male harbour porpoises. With these new basic parameters, indicators for endocrine disruptors can be developed in the future, aiming to detect how environmental factors could affect male fertility in wildlife.
T. Kesselring; S. Viquerat; L.L. Ijsseldijk; M. Langeheine; P. Wohlsein; A. Gröne; M. Bergmann; U. Siebert; R. Brehm. Testicular morphology and spermatogenesis in harbour porpoises (Phocoena phocoena). Theriogenology 2018, 126, 177 -186.
AMA StyleT. Kesselring, S. Viquerat, L.L. Ijsseldijk, M. Langeheine, P. Wohlsein, A. Gröne, M. Bergmann, U. Siebert, R. Brehm. Testicular morphology and spermatogenesis in harbour porpoises (Phocoena phocoena). Theriogenology. 2018; 126 ():177-186.
Chicago/Turabian StyleT. Kesselring; S. Viquerat; L.L. Ijsseldijk; M. Langeheine; P. Wohlsein; A. Gröne; M. Bergmann; U. Siebert; R. Brehm. 2018. "Testicular morphology and spermatogenesis in harbour porpoises (Phocoena phocoena)." Theriogenology 126, no. : 177-186.
Carolin Matschurat; Kristina Rode; Julia Hollenbach; Karola Wolf; Carola Urhausen; Andreas Beineke; Anne-Rose Günzel-Apel; Ralph Brehm. Impaired spermatogenesis, tubular wall disruption, altered blood-testis barrier composition and intratubular lymphocytes in an infertile Beagle dog - a putative case of autoimmune orchitis. Histol. Histopathol. 2018, 34, 525 -535.
AMA StyleCarolin Matschurat, Kristina Rode, Julia Hollenbach, Karola Wolf, Carola Urhausen, Andreas Beineke, Anne-Rose Günzel-Apel, Ralph Brehm. Impaired spermatogenesis, tubular wall disruption, altered blood-testis barrier composition and intratubular lymphocytes in an infertile Beagle dog - a putative case of autoimmune orchitis. Histol. Histopathol.. 2018; 34 (5):525-535.
Chicago/Turabian StyleCarolin Matschurat; Kristina Rode; Julia Hollenbach; Karola Wolf; Carola Urhausen; Andreas Beineke; Anne-Rose Günzel-Apel; Ralph Brehm. 2018. "Impaired spermatogenesis, tubular wall disruption, altered blood-testis barrier composition and intratubular lymphocytes in an infertile Beagle dog - a putative case of autoimmune orchitis." Histol. Histopathol. 34, no. 5: 525-535.
The current discussion concerning resource-efficient broiler production inevitably leads to diets with lowered crude protein (CP) levels. Therefore, the hypothesis was formed that crude protein reduction far below the recommended levels can significantly lower the nitrogen (N) content in litter, if essential amino acids are added and a constant lysine-arginine ratio is guaranteed. In a five-week feeding trial, 360 ROSS 308 broilers of both sexes were randomly assigned to four feeding groups with six replicates each with a standard three-phase feeding program (d 1–7, d 8–14, d 15–35). The control group was offered a complete diet with a common protein content found in practice (CP-% as fed; starter: 21.5, grower: 20.5, finisher: 20.0; lysine/arginine: 100/115). In the experimental diets the lysine/arginine ratio was constant, whereas the protein content was lowered in steps of 1.00 percent each with simultaneous supplementation of growth limiting amino acids. Feeding a diet with a 2.00 percent reduced protein content led to higher body weights after 34 days compared to the control (2329 g vs. 2192 g). The N content in the total litter decreased significantly with a 2.00 and 3.00 percent reduction in the CP content (51.2 vs. 46.2 or rather 36.2 g/kg dry matter (DM)). Meticulous balanced protein-reduced diets therefore allow a significant environmental relief.
Cristina Ullrich; Marion Langeheine; Ralph Brehm; Venja Taube; Diana Siebert; Christian Visscher. Influence of Reduced Protein Content in Complete Diets with a Consistent Arginine–Lysine Ratio on Performance and Nitrogen Excretion in Broilers. Sustainability 2018, 10, 3827 .
AMA StyleCristina Ullrich, Marion Langeheine, Ralph Brehm, Venja Taube, Diana Siebert, Christian Visscher. Influence of Reduced Protein Content in Complete Diets with a Consistent Arginine–Lysine Ratio on Performance and Nitrogen Excretion in Broilers. Sustainability. 2018; 10 (11):3827.
Chicago/Turabian StyleCristina Ullrich; Marion Langeheine; Ralph Brehm; Venja Taube; Diana Siebert; Christian Visscher. 2018. "Influence of Reduced Protein Content in Complete Diets with a Consistent Arginine–Lysine Ratio on Performance and Nitrogen Excretion in Broilers." Sustainability 10, no. 11: 3827.
For the reason that adult Sertoli cell specific connexin 43 knockout (SCCx43KO) mice show arrested spermatogenesis at spermatogonial level or Sertoli cell only tubules and significantly reduced germ cell (GC) numbers, the aims of the present study were (1) to characterize the remaining GC population and (2) to elucidate possible mechanisms of their fading. Apoptosis was analyzed in both, KO and wild type (WT) male littermates during postnatal development and in adulthood using TUNEL. Although GC numbers were significantly reduced in KO at 2 and 8 days postpartum (dpp) when compared to WT, no differences were found concerning apoptotic incidence between genotypes. From 10 dpp, the substantial GC deficiency became more obvious. However, significantly higher apoptotic GC numbers were seen in WT during this period, possibly related to the first wave of spermatogenesis, a known phenomenon in normal pubertal testes associated with increased apoptosis. Characterization of residual spermatogonia in postnatal to adult KO and WT mice was performed by immunohistochemical reaction against VASA (marker of GCs in general), Lin28 and Fox01 (markers for undifferentiated spermatogonia) and Stra8 (marker for differentiating spermatogonia and early spermatocytes). During puberty, the GC component in SCCx43KO mice consisted likely of undifferentiated spermatogonia, few differentiating spermatogonia and very few early spermatocytes, which seemed to be rapidly cleared by apoptosis. In adult KOs, spermatogenesis was arrested at the level of undifferentiated spermatogonia. Overall, our data indicate that Cx43 gap junctions in SCs influence male GC development and differentiation rather than their survival.
Kristina Rode; Karola Weider; Oliver Siegfried Damm; Joachim Wistuba; Marion Langeheine; Ralph Brehm. Loss of connexin 43 in Sertoli cells provokes postnatal spermatogonial arrest, reduced germ cell numbers and impaired spermatogenesis. Reproductive Biology 2018, 18, 456 -466.
AMA StyleKristina Rode, Karola Weider, Oliver Siegfried Damm, Joachim Wistuba, Marion Langeheine, Ralph Brehm. Loss of connexin 43 in Sertoli cells provokes postnatal spermatogonial arrest, reduced germ cell numbers and impaired spermatogenesis. Reproductive Biology. 2018; 18 (4):456-466.
Chicago/Turabian StyleKristina Rode; Karola Weider; Oliver Siegfried Damm; Joachim Wistuba; Marion Langeheine; Ralph Brehm. 2018. "Loss of connexin 43 in Sertoli cells provokes postnatal spermatogonial arrest, reduced germ cell numbers and impaired spermatogenesis." Reproductive Biology 18, no. 4: 456-466.
Connexin43 (Cx43) is the predominant testicular gap junction protein and in cases of impaired spermatogenesis, Cx43 expression has been shown to be altered in several mammals. Amongst other functions, Cx43 is supposed to regulate junction formation of the blood-testis barrier (BTB). The aim of the present study was to investigate the expression pattern of different tight junction (TJ) proteins of the murine BTB using SC-specific Cx43 knockout mice (SCCx43KO). Adult homozygous male SCCx43KO mice (SCCx43KO-/-) predominantly show an arrest of spermatogenesis and SC-only tubules that might have been caused by an altered BTB assembly, composition or regulation. TJ molecules claudin-3, -5 and -11 were examined in adult wild type (WT) and SCCx43KO-/- mice using immunohistochemistry (IHC) and quantitative real-time PCR (qRT-PCR). In this context, investigation of single tubules with residual spermatogenesis in SCCx43KO-/- mice was particularly interesting to identify a potential Cx43-independent influence of germ cells (GC) on BTB composition and dynamics. In tubules without residual spermatogenesis, a diffuse cytoplasmic distribution pattern for claudin-11 protein could be demonstrated in mutant mice. Nevertheless, claudin-11 seems to form functional TJ. Claudin-3 and -5 could not be detected immunohistochemically in the seminiferous epithelium of those tubules. Correspondingly, claudin-3 and -5 mRNA expression was decreased, providing evidence of generally impaired BTB dynamics in adult KO mice. Observations of tubules with residual spermatogenesis suggested a Cx43-independent regulation of TJ proteins by GC populations. To determine initial BTB formation in peripubertal SCCx43KO-/- mice, immunohistochemical staining and qRT-PCR of claudin-11 were carried out in adolescent SCCx43KO-/- and WT mice. Additionally, BTB integrity was functionally analysed using a hypertonic glucose fixative. These analyses revealed that SCCx43KO-/- mice formed an intact BTB during puberty in the same time period as WT mice, which however seemed to be accelerated.
Julia Hollenbach; Klaus Jung; Joanna Noelke; Hagen Gasse; Christiane Pfarrer; Mirja Koy; Ralph Brehm. Loss of connexin43 in murine Sertoli cells and its effect on blood-testis barrier formation and dynamics. PLOS ONE 2018, 13, e0198100 .
AMA StyleJulia Hollenbach, Klaus Jung, Joanna Noelke, Hagen Gasse, Christiane Pfarrer, Mirja Koy, Ralph Brehm. Loss of connexin43 in murine Sertoli cells and its effect on blood-testis barrier formation and dynamics. PLOS ONE. 2018; 13 (6):e0198100.
Chicago/Turabian StyleJulia Hollenbach; Klaus Jung; Joanna Noelke; Hagen Gasse; Christiane Pfarrer; Mirja Koy; Ralph Brehm. 2018. "Loss of connexin43 in murine Sertoli cells and its effect on blood-testis barrier formation and dynamics." PLOS ONE 13, no. 6: e0198100.
The gap junction protein connexin43 (CX43) plays a vital role in mammalian spermatogenesis by allowing for direct cytoplasmic communication between neighbouring testicular cells. In addition, different publications suggest that CX43 in Sertoli cells (SC) might be important for blood–testis barrier (BTB) formation and BTB homeostasis. Thus, through the use of the Cre-LoxP recombination system, a transgenic mouse line was developed in which only SC are deficient of the gap junction protein, alpha 1 (Gja1) gene.Gja1codes for the protein CX43. This transgenic mouse line has been commonly defined as the SC specific CX43 knockout (SCCx43KO) mouse line. Within the seminiferous tubule, SC aid in spermatogenesis by nurturing germ cells and help them to proliferate and mature. Owing to the absence of CX43 within the SC, homozygous KO mice are infertile, have reduced testis size, and mainly exhibit spermatogenesis arrest at the level of spermatogonia, seminiferous tubules containing only SC (SC-only syndrome) and intratubular SC-clusters. Although the SC specific KO of CX43 does not seem to have an adverse effect on BTB integrity, CX43 influences BTB composition as the expression pattern of different BTB proteins (like OCCLUDIN, β-CATENIN, N-CADHERIN, and CLAUDIN11) is altered in mutant males. The supposed roles of CX43 in dynamic BTB regulation, BTB assembly and/or disassembly and its possible interaction with other junctional proteins composing this unique barrier are discussed. Data collectively indicate that CX43 might represent an important regulator of dynamic BTB formation, composition and function.
Jonathan Gerber; Julia Heinrich; Ralph Brehm. Blood–testis barrier and Sertoli cell function: lessons from SCCx43KO mice. Reproduction 2016, 151, R15 -R27.
AMA StyleJonathan Gerber, Julia Heinrich, Ralph Brehm. Blood–testis barrier and Sertoli cell function: lessons from SCCx43KO mice. Reproduction. 2016; 151 (2):R15-R27.
Chicago/Turabian StyleJonathan Gerber; Julia Heinrich; Ralph Brehm. 2016. "Blood–testis barrier and Sertoli cell function: lessons from SCCx43KO mice." Reproduction 151, no. 2: R15-R27.
Testicular biopsy is still uncommonly used in equine andrological diagnostics although several studies supported that it is a relatively safe procedure. So far, no data exist regarding repeated testicular biopsies in stallions. In the present study, repeated testicular biopsy samples were obtained from four stallions at 4-week intervals alternatingly from both testes over the period of 1 year. The objectives were to assess (1) effects of repeated testicular biopsies by clinical, morphologic, histologic, ultrasonographic, and infrared thermographic examinations and (2) the utility of the biopsy samples for diagnostic purposes, namely hematoxylin–eosin staining, immunohistochemistry, Western blot analysis, and polymerase chain reaction. No significant side effects could be determined on clinical healthiness, testis size, libido, testicular blood flow, testicular histology, and scrotal surface temperature. The biopsy samples provided sufficient tissue for assessing spermatogenesis, detecting and localizing proteins (immunohistochemistry), and for protein/messenger RNA extraction for Western blot analyses and polymerase chain reaction. The authors conclude that taking even repeated testicular biopsies is a practicable and safe technique for equine practice having only minimal side effects. The biopsy specimens are suitable for various diagnostic cell and molecular biology applications for identifying testicular causes of male equine infertility. Prospectively, also therapeutic applications might be possible to extend the prospects of already established assisted reproductive techniques (e.g., testicular sperm extraction followed by intracytoplasmic sperm injection) in stallions suffering from obstructive azoospermia or oligospermia. Repeated testicular biopsies might also provide benefits for scientific work, for example, to monitor effects of pharmaceuticals or seasonal variations on testis function and spermatogenesis in the same animal.
Kristina Rode; Harald Sieme; Henning Otzen; Cornelia Schwennen; Matthias Lüpke; Peter Richterich; Rahel Schrimpf; Ottmar Distl; Ralph Brehm. Effects of Repeated Testicular Biopsies in Adult Warmblood Stallions and Their Diagnostic Potential. Journal of Equine Veterinary Science 2016, 38, 33 -47.
AMA StyleKristina Rode, Harald Sieme, Henning Otzen, Cornelia Schwennen, Matthias Lüpke, Peter Richterich, Rahel Schrimpf, Ottmar Distl, Ralph Brehm. Effects of Repeated Testicular Biopsies in Adult Warmblood Stallions and Their Diagnostic Potential. Journal of Equine Veterinary Science. 2016; 38 ():33-47.
Chicago/Turabian StyleKristina Rode; Harald Sieme; Henning Otzen; Cornelia Schwennen; Matthias Lüpke; Peter Richterich; Rahel Schrimpf; Ottmar Distl; Ralph Brehm. 2016. "Effects of Repeated Testicular Biopsies in Adult Warmblood Stallions and Their Diagnostic Potential." Journal of Equine Veterinary Science 38, no. : 33-47.
The formation of the blood-testis barrier (BTB) is defined as occurring with the first appearance of spermatocytes at around puberty and is vital for normal spermatogenesis. This barrier between two adjacent Sertoli cells (SCs) consists of a cell junctional protein complex, which includes tight junctions (TJs), adherens junctions, and gap junctions. In many mammalian species, BTB composition has already been investigated, whereas little is known about the equine BTB. In the present study, immunohistochemistry and qualitative Western Blot analysis were used to assess the expression and distribution patterns of the junctional proteins claudin-11 (TJ), zonula occludens-1 (TJ associated), N-cadherin (adherens junctions), and connexin 43 (gap junctions) in equine testes during tubular development and in testes of stallions exhibiting unilateral cryptorchidism. Therefore, testes of 21 warmblood stallions (aged 12 months-11 years) were obtained during routine surgical castration. In the normal adult equine testis, the junctional proteins are localized at the basolateral region of the seminiferous tubules forming a circumferential seal corresponding to the known BTB localization. N-cadherin is additionally expressed along the lateral SC surface. In immature seminiferous cords still lacking a lumen, a diffuse distribution pattern of the junctional proteins throughout the SC cytoplasm is visible. As lumen formation advances, the immunolocalization shifts progressively toward the basolateral SC membranes. Additionally, apoptotic germ cells were detected and quantified in prepubertal stallions using terminal deoxynucleotidyl transferase dUTP nick end labeling assay and correlated with junctional protein localization. In the retained testis of cryptorchid stallions, which exhibit an aberrant testicular morphology, a deviating expression of the junctional proteins is visible. The present data show for the first time that (1) the equine SC junctional complex contains claudin-11, zonula occludens-1, N-Cadherin, and connexin 43, as already described for men or mice, and that (2) different distribution patterns of these proteins exist during testicular development in the context of lumen formation (lumen scores: 1-7) and in retained testes of unilateral cryptorchid stallions.
K. Rode; H. Sieme; P. Richterich; R. Brehm. Characterization of the equine blood–testis barrier during tubular development in normal and cryptorchid stallions. Theriogenology 2015, 84, 763 -772.
AMA StyleK. Rode, H. Sieme, P. Richterich, R. Brehm. Characterization of the equine blood–testis barrier during tubular development in normal and cryptorchid stallions. Theriogenology. 2015; 84 (5):763-772.
Chicago/Turabian StyleK. Rode; H. Sieme; P. Richterich; R. Brehm. 2015. "Characterization of the equine blood–testis barrier during tubular development in normal and cryptorchid stallions." Theriogenology 84, no. 5: 763-772.
In vivo repopulation of decellularized allografts with recipient cells leads to a positive remodeling of the graft matrix in juvenile sheep. In light of the increasing number of heart valve replacements among older patients (>65 years), this study focused on the potential for matrix-guided tissue regeneration in elderly sheep. Pulmonary valve replacement was performed in seven-year old sheep using decellularized (DV), decellularized and CCN1-coated (RV), or decellularized and in vitro reendothelialized pulmonary allografts (REV) (n=6, each group). CCN1 coating was applied to support re-endothelialization. In vitro re-endothelialization was conducted with endothelial-like cells derived from peripheral blood. Echocardiograms of all grafts showed adequate graft function after implantation and at explantation 3 or 6 months later. All explants were macroscopically free of thrombi at explantation, and revealed repopulation of the allografts on the adventitial side of valvular walls and proximal in the cusps. Engrafted cells expressed vimentin, sm α-actin, and myosin heavy chain 2, while luminal cell lining was positive for vWF and eNOS. Cellular repopulation of valvular matrix demonstrates the capacity for matrix-guided regeneration even in elderly sheep but is not improved by in vitro endothelialization, confirming the suitability of decellularized matrix for heart valve replacement in older individuals.
Karolina Theodoridis; Igor Tudorache; Alexandru Calistru; Serghei Cebotari; Tanja Meyer; Samir Sarikouch; Christoph Bara; Ralph Brehm; Axel Haverich; Andres Hilfiker. Successful matrix guided tissue regeneration of decellularized pulmonary heart valve allografts in elderly sheep. Biomaterials 2015, 52, 221 -228.
AMA StyleKarolina Theodoridis, Igor Tudorache, Alexandru Calistru, Serghei Cebotari, Tanja Meyer, Samir Sarikouch, Christoph Bara, Ralph Brehm, Axel Haverich, Andres Hilfiker. Successful matrix guided tissue regeneration of decellularized pulmonary heart valve allografts in elderly sheep. Biomaterials. 2015; 52 ():221-228.
Chicago/Turabian StyleKarolina Theodoridis; Igor Tudorache; Alexandru Calistru; Serghei Cebotari; Tanja Meyer; Samir Sarikouch; Christoph Bara; Ralph Brehm; Axel Haverich; Andres Hilfiker. 2015. "Successful matrix guided tissue regeneration of decellularized pulmonary heart valve allografts in elderly sheep." Biomaterials 52, no. : 221-228.
The Sertoli cell (SC)-specific knockout (KO) of connexin43 (Cx43) results in spermatogenic arrest at the level of spermatogonia and/or SC-only syndrome. Histology of the interstitial compartment suggests Leydig cell (LC) hyperplasia. Our aim has been to investigate possible effects of the SC-specific KO of Cx43 (SCCx43KO) on interstitial LC. We therefore counted LC via the optical dissector method (per microliter of testicular tissue and per testis) and found LC to be significantly increased in SCCx43KO−/− compared with wild-type mice. Semiquantitative western blot together with Cx43 and 3β-hydroxysteroid dehydrogenase immunohistochemistry showed that Cx43 protein was significantly reduced and barely detectable in LC in adult SCCx43KO−/− mice. This reduction of Cx43 protein was accompanied by a reduction of Cx43 mRNA as analyzed by laser-assisted microdissection of interstitial cells and subsequent quantitative real-time polymerase chain reaction (PCR). Interestingly, Cx45, another recently detected connexin in LC, was also downregulated. Preliminary qualitative data of LC differentiation markers (Thb2, Hsd3b6) and a steroidogenic marker (Hsd17b3) obtained by reverse transcription plus PCR revealed no obvious differences. Thus, the loss of Cx43 in SC also provokes the downregulation of connexins in interstitial LC at the transcriptional and translational levels. Moreover, SCCx43KO leads to alterations in LC numbers. Despite these alterations, steroidogenesis seems not to be impaired. Further studies, including ultrastructural analysis of the tissue as well as quantitative examination of additional LC markers and testosterone, and functional in vitro experiments, should provide more information about LC differentiation and function in SCCx43KO−/− mice.
Joanna Noelke; Joachim Wistuba; Oliver S. Damm; Daniela Fietz; Jonathan Gerber; Marion Gaehle; Ralph Brehm. A Sertoli cell-specific connexin43 knockout leads to altered interstitial connexin expression and increased Leydig cell numbers. Cell and Tissue Research 2015, 361, 633 -644.
AMA StyleJoanna Noelke, Joachim Wistuba, Oliver S. Damm, Daniela Fietz, Jonathan Gerber, Marion Gaehle, Ralph Brehm. A Sertoli cell-specific connexin43 knockout leads to altered interstitial connexin expression and increased Leydig cell numbers. Cell and Tissue Research. 2015; 361 (2):633-644.
Chicago/Turabian StyleJoanna Noelke; Joachim Wistuba; Oliver S. Damm; Daniela Fietz; Jonathan Gerber; Marion Gaehle; Ralph Brehm. 2015. "A Sertoli cell-specific connexin43 knockout leads to altered interstitial connexin expression and increased Leydig cell numbers." Cell and Tissue Research 361, no. 2: 633-644.