Differential phosphorylation of two serine clusters in mouse HORMAD1 during meiotic prophase I progression.

Mouse HORMAD1 is a phospho-protein involved in multiple functions during meiotic prophase I. To obtain insight into the significance of its phosphorylation, we generated phospho-specific antibodies against two serine residues, Ser307 and Ser378, representing each of two serine clusters in mouse HORMAD1. The Ser307 phosphorylation is detectable from early leptotene substage in both wild-type and Spo11-/- spermatocytes, indicating that Ser307 is a primary and SPO11-independent phosphorylation site. In contrast, the Ser378 phosphorylation is negligible at earlier substages in wild-type and Spo11-/- spermatocytes. After mid-zygotene substage, the Ser378 phosphorylation is abundant on unsynapsed chromosome axes in wild-type spermatocytes and is detected only in a part of unsynapsed chromosome axes in Spo11-/- spermatocytes. We also generated a non-phosphorylated Ser307-specific antibody and found that Ser307 is phosphorylated on sex chromosome axes but is almost entirely unphosphorylated on desynapsed chromosome axes in diplotene spermatocytes. These results demonstrated a substage-specific phosphorylation status of mouse HORMAD1, which might be associated with multiple substage-specific functions.

Aquaporin-5 Protein Is Selectively Reduced in Rat Parotid Glands under Conditions of Fasting or a Liquid Diet.

Aquaporin-5 (AQP5) water channel, transmembrane protein 16A (TMEM16A) Ca2+-activated Cl- channel, and Na+-K+-2Cl- cotransporter (NKCC1) are membrane proteins on salivary gland acinar cells that function in watery saliva secretion. We examined their expression changes in rat parotid glands under reduced mastication. Rats were either fed regular chow as a control group, fasted for 48 hr or fed a liquid diet for 48 hr or 1 week to reduce mastication. The parotid glands were then resected to analyze the protein and mRNA levels by immunofluorescence, immunoblotting, and reverse-transcription quantitative PCR (RT-qPCR). AQP5 protein was significantly decreased in both liquid diet groups and the fasting group but its mRNA levels showed no apparent changes compared with the control group. The protein and mRNA levels of TMEM16A and NKCC1 showed no significant changes between any of the groups other than an increase in NKCC1 mRNA in the 1-week liquid diet group. These results suggest that reduced mastication may increase the AQP5 protein degradation, but not that of other membrane proteins necessary for saliva secretion.

Progress in the Development of Stem Cell-Derived Cell-Free Therapies for Skin Aging.

Introduction: The skin is a vital organ as the body's largest barrier, but its function declines with aging. Therefore, research into effective regeneration treatments must continue to advance. Stem cell transplantation, a cell-based therapy, has become a popular skin-aging treatment, although it comes with drawbacks like host immune reactions. Stem cell-derived cell-free therapies have emerged as an alternative, backed by promising preclinical findings. Stem cell secretomes and extracellular vesicles (EVs) are the key components in cell-free therapy from stem cells. However, comprehensive reviews on the mechanisms of such treatments for skin aging are still limited. Purpose: This review discusses stem cell-derived cell-free therapy's potential mechanisms of action related to skin aging prevention by identifying specific molecular targets suitable for the interventions. Methods: A search identified 27 relevant in vitro studies on stem cell-derived cell-free therapy interventions in skin aging model cells without restricting publication years using PubMed, Scopus, and Google Scholar. Results: Stem cell-derived cell-free therapy can prevent skin aging through various mechanisms, such as (1) involvement of multiple regenerative pathways [NFkb, AP-1, MAPK, P-AKT, NRF2, SIRT-1]; (2) oxidative stress regulation [by reducing oxidants (HO-1, NQO1) and enhancing antioxidants (SOD1, CAT, GP, FRAP)]; (3) preventing ECM degradation [by increasing elastin, collagen, HA, TIMP, and reducing MMP]; (4) regulating cell activity [by reducing cell senescence (SA-ß-gal), apoptosis, and cell cycle arrest (P53, P12, P16); and enhancing autophagy, cell migration, and cell proliferation (Ki67)] (5) Regulating the inflammatory pathway [by reducing IL-6, IL-1, TNF-⍺, and increasing TGF-ß]. Several clinical trials have also revealed improvements in wrinkles, elasticity, hydration, pores, and pigmentation. Conclusion: Stem cell-derived cell-free therapy is a potential novel treatment for skin aging by cell rejuvenation through various molecular mechanisms.

Loss of VAMP5 in mice results in duplication of the ureter and insufficient expansion of the lung.

BACKGROUND: Vesicle-associated membrane protein 5 (VAMP5) is a member of the SNARE protein family, which regulates the docking and fusion of membrane vesicles within cells. Previously, we reported ubiquitous expression of VAMP5 proteins in various organs except the brain and small intestine. However, the precise roles of VAMP5 in each organ remain unclear. To explore the roles of VAMP5 in vivo, we generated VAMP5 knockout (KO) mice. RESULTS: VAMP5 KO mice showed low birth rate and low body weight. KO embryos grew normally in the uterus, and tended to die around birth. Anatomical analysis revealed that viable KO mice often exhibited duplication of the ureter, and dead KO mice showed insufficient expansion of the lung. VAMP5 was localized in the epithelial cells of the ureter and terminal bronchiole. CONCLUSIONS: VAMP5 KO mice showed a low birth rate and abnormalities of the urinary and respiratory systems. VAMP5 KO mice died around birth, possibly due to defects in vesicoureteral flow and breathing. The results presented could provide a basis for future studies to understand the roles of VAMP5 protein. Developmental Dynamics 247:754-762, 2018. © 2018 Wiley Periodicals, Inc.

A novel imaging method for correlating 2D light microscopic data and 3D volume data based on block-face imaging.

We have developed an imaging method designated as correlative light microscopy and block-face imaging (CoMBI), which contributes to improve the reliability of morphological analyses. This method can collect both the frozen sections and serial block-face images in a single specimen. The frozen section can be used for conventional light microscopic analysis to obtain 2-dimensional (2D) anatomical and molecular information, while serial block-face images can be used as 3-dimensional (3D) volume data for anatomical analysis. Thus, the sections maintain positional information in the specimen, and allows the correlation of 2D microscopic data and 3D volume data in a single specimen. The subjects can vary in size and type, and can cover most specimens encountered in biology. In addition, the required system for our method is characterized by cost-effectiveness. Here, we demonstrated the utility of CoMBI using specimens ranging in size from several millimeters to several centimeters, i.e., mouse embryos, human brainstem samples, and stag beetle larvae, and present successful correlation between the 2D light microscopic images and 3D volume data in a single specimen.