High percentage of midpiece defects in Brangus bull sperm with no reduction in sperm kinematics.

The study investigated midpiece defects in sperm from a 5-year-old Brangus bull with a high rate of semen batch rejection, due to morphologically abnormal sperm, with no reduction in sperm kinematics. A comprehensive evaluation was conducted over a 16-month period, involving 28 ejaculates. Notably, despite the high proportion of midpiece defects (average 37.73%, from 3% to 58%), the study revealed stable sperm production, with no discernible differences in the kinematic data before and after cryopreservation. Electron microscopy identified discontinuities in the mitochondrial sheath, characteristic of midpiece aplasia (MPA). The anomalies were attributed to be of genetic origin, as other predisposing factors were absent. Additionally, the electron microscopy unveiled plasma membrane defects, vacuoles and chromatin decondensation, consistent with previous findings linking acrosome abnormalities with midpiece defects. The findings underscored the necessity of conducting thorough laboratory evaluations before releasing cryopreserved semen for commercialization. Despite substantial morphological alterations, the initial semen evaluation data indicated acceptable levels of sperm kinematics, emphasizing the resilience of sperm production to severe morphological changes. This case report serves as a contribution to the understanding of midpiece defects in bull sperm, emphasizing the need for meticulous evaluation and quality control in semen processing and commercialization.

Senescence on Dental Pulp Cells: Effects on Morphology, Migration, Proliferation, and Immune Response.

INTRODUCTION: Evidence indicates that senescence can affect essential dental pulp functions, such as defense capacity and repair, consequently affecting the successes of conservative endodontic treatments. This study aims to evaluate the effects of senescence on the morphology, migration, proliferation, and immune response of human dental pulp cells. METHODS: Cells were treated with doxorubicin to induce senescence, confirmed by ß-galactosidase staining. Morphological changes, cellular proliferation, and migration were evaluated by scanning electron microscopy, trypan blue cells, and the scratch method, respectively. Modifications in the immune response were evaluated by measuring the genes for pro-inflammatory cytokines tumor necrosis factor alpha and interleukin (IL)-6 and anti-inflammatory cytokines transforming growth factor beta 1 and IL-10 using the real time polymerase chain reaction assay. RESULTS: An increase in cell size and a decrease in the number of extensions were observed in senescent cells. A reduction in the proliferative and migratory capacity was also found in senescent cells. In addition, there was an increase in the gene expression of inflammatory cytokines tumor necrosis factor alpha and IL-6 and a decrease in the gene expression of IL-10 and transforming growth factor beta-1, suggesting an exacerbated inflammatory situation associated with immunosuppression. CONCLUSIONS: Cellular senescence is possibly a condition that affects prognoses of conservative endodontic treatments, as it affects primordial cellular functions related to this treatment.

Antileishmanial activity of 2-amino-thiophene derivative SB-200.

Leishmaniasis, presenting the highest number of cases worldwide is one of the most serious Neglected Tropical Diseases (NTDs). Clinical manifestations are intrinsically related to the host's immune response making immunomodulatory substances the target of numerous studies on antileishmanial activity. The currently available drugs used for treatment present various problems including high toxicity, low efficacy, and associated drug resistance. The search for therapeutic alternatives is urgent, and in this context, thiophene derivatives appear to be a promising therapeutic alternative (many have shown promising anti-leishmanial activity). The objective of this study was to investigate the antileishmanial activity of the 2-amino-thiophenic derivative SB-200. The thiophenic derivative was effective in inhibiting the growth of Leishmania braziliensis, Leishmania major, and Leishmania infantum promastigotes, obtaining respective IC50 values of 4.25 µM, 4.65 µM, and 3.96 µM. For L. infantum, it was demonstrated that the antipromastigote effect of SB-200 is associated with cell membrane integrity losses, and with morphological changes observed during scanning and transmission electron microscopy. Cytotoxicity was performed for J774.A1 macrophages and VERO cells, to obtain a CC50 of 42.52 µM and a SI of 10.74 for macrophages and a CC50 of 39.2 µM and an SI of 9.89 for VERO cells. The anti-amastigote activity of SB-200 revealed an IC50 of 2.85 µM and an SI of 14.97 against macrophages and SI of 13.8 for VERO cells. The anti-amastigote activity of SB-200 is associated with in vitro immunomodulation. For acute toxicity, SB-200 against Zophobas morio larvae permitted 100% survival. We conclude that the 2-amino-thiophenic derivative SB-200 is a promising candidate for in vivo anti-leishmania drug tests to evaluate its activity, efficacy, and safety.

Host defense peptides combined with MTA extract increase the repair in dental pulp cells: in vitro and ex vivo study.

Host Defense Peptides (HDPs) have, in previous studies, been demonstrating antimicrobial, anti-inflammatory, and immunomodulatory capacity, important factors in the repair process. Knowing these characteristics, this article aims to evaluate the potential of HDPs IDR1018 and DJK-6 associated with MTA extract in the repair process of human pulp cells. Antibacterial activity of HDPs, MTA and HDPs combined with MTA in Streptococcus mutans planktonic bacteria and antibiofilm activity was evaluated. Cell toxicity was assayed with MTT and cell morphology was observed by scanning electron microscopy (SEM). Proliferation and migration of pulp cells were evaluated by trypan blue and wound healing assay. Inflammatory and mineralization related genes were evaluated by qPCR (IL-6, TNFRSF, DSPP, TGF-ß). Alkaline phosphatase, phosphate quantification and alizarin red staining were also verified. The assays were performed in technical and biological triplicate (n = 9). Results were submitted for the calculation of the mean and standard deviation. Then, normality verification by Kolmogorov Smirnov test, analyzing one-way ANOVA. Analyses were considered at a 95% significance level, with a p-value < 0.05. Our study demonstrated that HDPs combined with MTA were able to reduce biofilms performed in 24 h and biofilm performed over 7 days S. mutans biofilm (p < 0.05). IDR1018 and MTA, as well as their combination, down-regulated IL-6 expression (p < 0.05). Tested materials were not cytotoxic to pulp cells. IDR1018 induced high cell proliferation and combined with MTA induced high cellular migration rates in 48 h (p < 0.05). Furthermore, the combination of IDR1018 and MTA also induced high expression levels of DSPP, ALP activity, and the production of calcification nodules. So, IDR-1018 and its combination with MTA could assist in pulp-dentine complex repair process in vitro.

Regulatory T-Cell Enhancement, Expression of Adhesion Molecules, and Production of Anti-Inflammatory Factors Are Differentially Modulated by Spheroid-Cultured Mesenchymal Stem Cells.

The culture of mesenchymal stem cells (MSCs) as spheroids promotes a more physiological cellular behavior, as it more accurately reflects the biological microenvironment. Nevertheless, mixed results have been found regarding the immunosuppressive properties of spheroid-cultured MSCs (3D-MSCs), the mechanisms of immunoregulation of 3D-MSCs being scarcely described at this point. In the present study, we constructed spheroids from MSCs and compared their immunosuppressive potential with that of MSCs cultured in monolayer (2D-MSCs). First, we evaluated the ability of 2D-MSCs and 3D-MSCs to control the activation and proliferation of T-cells. Next, we evaluated the percentage of regulatory T-cells (Tregs) after the co-culturing of peripheral blood mononuclear cells (PBMCs) with 2D-MSCs and 3D-MSCs. Finally, we investigated the expression of adhesion molecules, as well as the expressions of several anti-inflammatory transcripts in 2D-MSCs and 3D-MSCs maintained in both inflammatory and non-inflammatory conditions. Interestingly, our data show that several anti-inflammatory genes are up-regulated in 3D-MSCs, and that these cells can control T-cell proliferation. Nevertheless, 2D-MSCs are more efficient in suppressing the immune cell proliferation. Importantly, contrary to what was observed in 3D-MSCs, the expressions of ICAM-1 and VCAM-1 are significantly upregulated in 2D-MSCs exposed to an inflammatory environment. Furthermore, only 2D-MSCs are able to promote the enhancement of Tregs. Taken together, our data clearly show that the immunosuppressive potential of MSCs is significantly impacted by their shape, and highlights the important role of cell-cell adhesion molecules for optimal MSC immunomodulatory function.