The role of Rho-associated kinase inhibitor, Y-27632 on primary culture of ovine spermatogonial stem cells

The access to sufficient numbers of spermatogonial stem cells (SSCs) is a prerequisite for the study of their regulation and further biomanipulation. Rho kinase (ROCK) belongs to a family of serine/threonine kinases and involves in a wide range of fundamental cellular functions. The aim of the present study was to study the effect of ROCK inhibitor, Y-27632 (0.1-40 µM), during the primary culture of ovine SSCs. SSCs were collected from 3-5-month-olds lamb testes. The viability of SSCs, the apoptosis assay of SSCs, the intracellular reactive oxygen species (ROS) analysis, and the SSCs markers and apoptosis-related gene expressions were detected by MTT reduction assay, Annexin V–FITC/ Propidium Iodide (PI) dual staining, flow cytometry and real-time-PCR studies, respectively. Morphological analyses indicated that the 5-10 µM Y-27632 had an optimal effect on the number of presumptive SSCs colonies and the area covered by them after a 10 days culture. The cell viability, apoptosis and necrosis of SSCs after 10 days culture were not affected in comparison with the control group, and the 20 µM of Y-27632 resulted in significantly decreased cell viability (P<0.05) and an increased necrosis of cells. On day 10 after culture, the expression of P53 was decreased with an increase from 0 to 10 µM in the Y-27632 dose. In the 20 µM Y-27632 group, the expressions of P53 and Bax were higher and the Bcl-2 was lower than other groups and these values were significantly different from 5 and 10 µM Y-27632 groups (P<0.05). The level of intracellular ROS was decreased with an increase in the Y-27632 dose from 5 to 20 µM in comparison with the control group. In conclusion, the present study demonstrated that Y-27632 at a concentration of 5-10 µM provided optimal culture conditions for the primary culture of ovine SSCs.(AU)

Lactobacillus acidophilus reduces Listeria monocytogenes infection by inhibiting mitogen-activated protein kinase genes in growing rabbits

We aimed to investigate the effect of Lactobacillus acidophilus ACCC11073 on the growth performance, oxidation, inflammation, and mitogen-activated protein kinase (MAPK) family genes of rabbits infected with Listeria monocytogenes (L. monocytogenes) using antibiotic enrofloxacin hydrochloride (EH) as a reference. There were four treatments including negative control, positive control with L. monocytogenes infection on the first day of feeding trial (PC), PC + EH at 40 mg kg−1, and PC + L. acidophilus at 108 CFU kg−1 of diet using 240 weaned growing rabbits. The results showed that L. monocytogenes infection worsened growth performance of rabbits, whereas EH or L. acidophilus supplementation partially recovered body weight gain, but did not reach the levels of the negative control. Listeria acidophilus and EH decreased L. monocytogenes loads in caecum, liver, spleen, and lymph node, serum oxidative markers including diamine oxidase, malondialdehyde, and protein carbonyl, serum IL-1ß, IL-6, and TNF-α. The decreased effects of EH on IL-1ß and TNF-α were more pronounced than that of the probiotic. Treatments EH and probiotic also de-regulated the mRNA levels of MAPK1, 3, 6, and 14. Listeria acidophilus exhibits a similar effect to EH against L. monocytogenes in rabbits, and the regulation on inflammatory process is via MAPK family genes. The results suggest that L. acidophilus can be used as a feed additive against L. monocytogenes infection.(AU)

Oclacitinib em processos dermatológicos da medicina veterinária - revisão de literatura / Oclacitinib in dermatological disease processes in veterinary medicine - a literature review / El oclacitinib para cuadros dermatológicos en medicina veterinaria - revisión de literatura

Muitas citocinas, incluindo as interleucinas (ILs), os interferons (IFNs) e outras moléculas, utilizam a via de transdução e transcrição Janus-kinase (Janus-kinase/signal transducers activators of transcription – JAK-STAT) para a transmissão de sinais da membrana citoplamástica para o núcleo das células. Essas citocinas inflamatórias dependem dessa sinalização JAK-STAT, que é indispensável para a função imune e hematopoiética. Sendo assim, desenvolveram-se inibidores de JAKs para o tratamento de diversas doenças inflamatórias autoimunes e hematológicas em seres humanos. Os inibidores de JAKs têm se mostrado eficazes no tratamento de dermatite atópica, alopecia areata, psoríase e vitiligo em seres humanos. Dentre os inibidores de JAK incluem-se: tofacitinib, ruxolitinib, baricitinib e oclacinitib. O oclacinitib é de uso exclusivo da medicina veterinária, e estudos recentes demonstram que esse fármaco tem um efeito rápido na redução do prurido e nas lesões de cães com dermatite atópica. Este trabalho teve como objetivo revisar essa nova e promissora classe de medicamentos, com foco no oclacinitib usado na medicina veterinária.

The Janus kinase signal transducer and activator of transcription (JAK-STAT) pathway is utilized by cytokines, including interleukins, interferons (IFNs), and other molecules to transmit signals from the cell membrane to the nucleus. Inflammatory cytokines rely on JAK-STAT signaling, which is indispensable for immune and hematopoietic function. JAK inhibitors were developed for the treatment of a variety of inflammatory, immune-mediated, and hematopoietic disorders in humans. Evidence suggests that JAK inhibitors are efficacious in the treatment of atopic dermatitis, alopecia areata, psoriasis, and vitiligo. The first generation of JAK inhibitors includes tofacitinib, ruxolitinib, baricitinib, and oclacitinib. Oclacitinib is used exclusively in veterinary medicine, and recent studies have shown that this drug has a rapid effect on pruritus and lesion reduction in dogs with atopic dermatitis. This article aimed to review this new class of drugs, focusing on the use of oclacitinib in veterinary medicine.

Muchas citocinas, entre ellas las interleucinas (ILs), interferones (IFNs) y otras moléculas utilizan la vía de la transducción y transcripción Janus-quinasa (Janus kinase/signal transducers and activators oftranscription - JAK-STAT) para la transmisión de señales desde la membrana citoplasmática hacia el núcleo celular. Esas citocinas inflamatorias dependen de esa señalización JAK-STAT, indispensable para la función inmune y hematopoyética. En seres humanos, se han desarrollado inhibidores de JAKs para el tratamiento de diferentes tipos de enfermedades inflamatorias autoinmunes y hematológicas. También en humanos, los inhibidores de JAKs se han mostrado eficientes para el tratamiento de dermatitis atópica, alopecia areata, psoriasis y vitíligo. Algunos de estos inhibidores son el tofacitinib, ruxolitinib, baricitinib y el oclacitinib. Esta última droga, oclacitinib, es de uso exclusivo en medicina veterinaria y estudios recientes han demostrado que tiene rápido efecto en la disminución deli prurito y lesiones de perros con dermatitis atópica. Este trabajo tuvo como objetivo revisar la literatura relacionada con esta nueva y prometedora clase de medicamentos, haciendo foco en el oclacitinib utilizado en medicina veterinaria.

Oclacitinib em processos dermatológicos da medicina veterinária - revisão de literatura / Oclacitinib in dermatological disease processes in veterinary medicine - a literature review / El oclacitinib para cuadros dermatológicos en medicina veterinaria - revisión de literatura

Muitas citocinas, incluindo as interleucinas (ILs), os interferons (IFNs) e outras moléculas, utilizam a via de transdução e transcrição Janus-kinase (Janus-kinase/signal transducers activators of transcription JAK-STAT) para a transmissão de sinais da membrana citoplamástica para o núcleo das células. Essas citocinas inflamatórias dependem dessa sinalização JAK-STAT, que é indispensável para a função imune e hematopoiética. Sendo assim, desenvolveram-se inibidores de JAKs para o tratamento de diversas doenças inflamatórias autoimunes e hematológicas em seres humanos. Os inibidores de JAKs têm se mostrado eficazes no tratamento de dermatite atópica, alopecia areata, psoríase e vitiligo em seres humanos. Dentre os inibidores de JAK incluem-se: tofacitinib, ruxolitinib, baricitinib e oclacinitib. O oclacinitib é de uso exclusivo da medicina veterinária, e estudos recentes demonstram que esse fármaco tem um efeito rápido na redução do prurido e nas lesões de cães com dermatite atópica. Este trabalho teve como objetivo revisar essa nova e promissora classe de medicamentos, com foco no oclacinitib usado na medicina veterinária.(AU)

The Janus kinase signal transducer and activator of transcription (JAK-STAT) pathway is utilized by cytokines, including interleukins, interferons (IFNs), and other molecules to transmit signals from the cell membrane to the nucleus. Inflammatory cytokines rely on JAK-STAT signaling, which is indispensable for immune and hematopoietic function. JAK inhibitors were developed for the treatment of a variety of inflammatory, immune-mediated, and hematopoietic disorders in humans. Evidence suggests that JAK inhibitors are efficacious in the treatment of atopic dermatitis, alopecia areata, psoriasis, and vitiligo. The first generation of JAK inhibitors includes tofacitinib, ruxolitinib, baricitinib, and oclacitinib. Oclacitinib is used exclusively in veterinary medicine, and recent studies have shown that this drug has a rapid effect on pruritus and lesion reduction in dogs with atopic dermatitis. This article aimed to review this new class of drugs, focusing on the use of oclacitinib in veterinary medicine.(AU)

Muchas citocinas, entre ellas las interleucinas (ILs), interferones (IFNs) y otras moléculas utilizan la vía de la transducción y transcripción Janus-quinasa (Janus kinase/signal transducers and activators oftranscription - JAK-STAT) para la transmisión de señales desde la membrana citoplasmática hacia el núcleo celular. Esas citocinas inflamatorias dependen de esa señalización JAK-STAT, indispensable para la función inmune y hematopoyética. En seres humanos, se han desarrollado inhibidores de JAKs para el tratamiento de diferentes tipos de enfermedades inflamatorias autoinmunes y hematológicas. También en humanos, los inhibidores de JAKs se han mostrado eficientes para el tratamiento de dermatitis atópica, alopecia areata, psoriasis y vitíligo. Algunos de estos inhibidores son el tofacitinib, ruxolitinib, baricitinib y el oclacitinib. Esta última droga, oclacitinib, es de uso exclusivo en medicina veterinaria y estudios recientes han demostrado que tiene rápido efecto en la disminución deli prurito y lesiones de perros con dermatitis atópica. Este trabajo tuvo como objetivo revisar la literatura relacionada con esta nueva y prometedora clase de medicamentos, haciendo foco en el oclacitinib utilizado en medicina veterinaria.(AU)

MCM3 could be a better marker than Ki-67 for evaluation of dysplastic oral lesions: an immunohistochemical study.

BACKGROUND: The aim of this study was to identify the expression of MCM3, Ki-67 and p27 in normal mucosa, leucoplakia and oral squamous cell carcinoma (OSCC) and determine whether altered expression could serve as a prognostic marker of a malignant progression of dysplastic lesions. METHODS: The samples were collected from 37 patients with oral leucoplakia (13 with mild dysplasia - MLD, 12 with moderate dysplasia - MD and 12 with severe dysplasia - SD). Eleven samples of mouth floor mucocele (M) and 50 floor mouth and tongue samples OSCC of untreated patients were included in this study. Immunohistochemical expression of MCM3, Ki-67 and p27 of all the groups was analysed. Kruskal-Wallis and Dunn's test were used to determine differences among groups, and a Pearson's correlation test was used to evaluate the correlation between the proteins. RESULTS: Ki-67 expression was higher in OSCC than M (P < 0.001) and MLD (P < 0.01) groups, and there was a lower expression in M compared with MD and SD (P < 0.05). Regarding p27, its expression was lower in OSCC compared with M, MD and SD. MCM3 expression was lower in M compared with SD and OSCC (P < 0.001), and MLD showed a lower expression when compared SD (P < 0.01) and OSCC (P < 0.001). Moreover, a better correlation was observed between the proteins MCM3 and p27 than between Ki-67 and p27 proteins when all lesions were examined together. CONCLUSIONS: This study showed that MCM3 could be a better marker than Ki-67 for evaluation of dysplastic oral lesions.

Homology modeling of LmxMPK4 of Leishmania mexicana and virtual screening of potent inhibitors against it.

Leishmaniasis is one of the most important diseases of mankind. In the life cycle of Leishmania mexicana, two most important developmental stages are observed. In insect vector it is in promastigote form and in mammalian macrophages is the amastigote form. The family of protein kinases are extremely important regulators of many different cellular processes such as transcriptional control, cell cycle development and differentiation, and also draw much attention as possible drug targets for protozaon parasites. Leishmania mexicana mitogen activated protein kinase 4 (LmxMPK4) is essential for proliferation and survival of the parasite promastigote and amastigote forms and is a potential drug target for leishmaniasis. The existing therapy for leishmaniasis is not enough due to host toxicity and drug resistance. The experimental 3D structure of this protein has not yet been determined. In this study, we have used homology modelling techniques to generate the 3D structure of LmxMPK4 and selected effective inhibitors by ZINC database on the basis of structure of berberine alkaloid for molecular docking studies with LmxMPK4. The inhibitors ZINC05999210, ZINC40402312 and ZINC40977377 were found to be more potent for inhibition of leishmaniasis due to the robust binding affinity and strong inhibition constant (Ki) of the protein-ligand interactions. This finding may help to understand the nature of MAP kinase and development of specific anti-leishmanial therapies.

Arginine vasopressin controls p27Kip1 protein expression by PKC activation andirreversibly inhibits the proliferation of K-Ras-dependent mouse Y1 adrenocortical malignant cells

The neurohypophyseal hormone arginine vasopressin (AVP) is a classic mitogen in many cells. In K-Rasdependent mouse Y1 adrenocortical malignant cells, AVP elicits antagonistic responses such as the activation of the PKC and the ERK1/2 mitogenic pathways to down-regulate cyclin D1 gene expression, which induces senescence-associated â-galactosidase (SA-âGal) and leads to cell cycle arrest. Here, we report that in the metabolic background of Y1 cells, PKC activation either by AVP or by PMA inhibits the PI3K/Akt pathway and stabilises the p27Kip1 protein even in the presence of the mitogen fibroblast growth factor 2 (FGF2). These results suggest that p27Kip1 is a critical signalling node in the mechanisms underlying the survival of the Y1 cells. In Y1 cells that transiently express wild-type p27Kip1, AVP caused a severe reduction in cell survival, as shown by clonogenic assays. However, AVP promoted the survival of Y1 cells transiently expressing mutant p27-S10A or mutant p27-T187A, which cannot be phosphorylated at Ser10 and Thr187, respectively. In addition, PKC activation by PMA mimics the toxic effect caused by AVP in Y1 cells, and inhibition of PKC completely abolishes the effects caused by both PMA and AVP in clonogenic assays. The vulnerability of Y1 cells during PKC activation is a phenotype conditioned upon K-ras oncogene amplification because K-Ras down-regulation with an inducible form of the dominant-negative mutant H-RasN17 has resulted in Y1 cells that are resistant to AVP's deleterious effects. These data show that the survival destabilisation of K-Rasdependent Y1 malignant cells by AVP requires large quantities of the p27Kip1 protein as well as phosphorylation of the p27Kip1 protein at both Ser10 and Thr187.