Oxytocin, and its teleost homolog isotocin, are neuropeptides that regulate social affiliation and cohesion across vertebrates. The African cichlid Astatotilapia burtoni is a powerful model for studying the neuroendocrine basis of social behavior due to its dynamic hierarchies and behavioral plasticity. While oxytocin signaling is implicated in adult social behaviors, its role in early social cohesion remains unclear. This study tests whether isotocin mediates the development of affiliative behavior in sexually immature juveniles by examining shoaling behavior in cichlids with and without functional signaling. Juveniles from CRISPR-Cas9 isotocin-knockout heterozygous crosses were genotyped via fin clips, PCR, and gel electrophoresis to identify wild type (WT), heterozygous (Het), and knockout (KO) individuals. Shoaling was assessed in petri dish arenas under standardized conditions. Groups of six same-genotype juveniles were acclimated for 30 minutes and recorded for 45 minutes, with behavior quantified using Noldus EthoVision tracking software. Welch’s t-tests revealed that KO fish showed significantly increased locomotor activity (distance and velocity, p = 0.002) and reduced social cohesion, including greater inter-individual distance (p = 0.002) and less physical contact (p = 0.009), compared to WT. Het individuals displayed intermediate behavior. These findings suggest oxytocin signaling is critical for early social cohesion, highlighting conserved mechanisms underlying the developmental origins of social behavior.

The thermogenic properties of brown adipose tissue (BAT) make it a promising area of study in combating metabolic disease. Amino-terminal enhancer of split (AES) is a member of the transducin-like enhancer of split (TLE) family which lacks the WD-repeat (WDR) domain classically responsible for recruiting transcription factors. The mechanism of AES in regulating transcription in brown adipocytes has yet to be established. Based on its unique truncated nature, we hypothesize that AES forms heterotetramers with TLE3 which enable gene regulation in brown adipocytes. In this investigation, AlphaFold2-Multimer and HADDOCK were used to produce a predictive structural model of the AES-TLE3 heterodimer. In silico alanine scanning revealed that the modeled complex is stabilized primarily by a central hydrophobic patch while retaining canonical TLE coiled-coil interface geometry. To validate this model, we generated stable cell lines expressing interface-disrupting AES mutants which have been validated for heterodimer disruption in concordance with computational predictions via coimmunoprecipitation. Our model suggests that repression of full-length TLE members by AES occurs via formation of nonfunctional heterotetramers. In the brown adipocyte, the promotion of white adipocyte development by TLE3 suggests that this dominant-negative effect of AES contributes to thermogenic development. This model provides a structural framework for elucidating the mechanism of AES in promoting thermogenic gene expression in brown adipocytes.

High-resolution micro-computed tomography (Micro-CT) is an X-ray imaging modality for visualizing complex internal bone structures in 3D. However, because Micro-CT scanners have a small field of view, specimens often need to be reduced in size, a process that is especially challenging when fragile anatomical features are embedded within dense bone. This protocol offers an alternative to manual sectioning by using a laser cutter guided by low-resolution CT imaging. This study included three cochlea samples embedded in human temporal bones, each varying in morphology and clinical condition. Low-resolution CT scans were used to map the position of the cochlea, guiding alignment and sectioning with a CO₂ laser cutter. The prepared specimens were then imaged using high-resolution Micro-CT and analyzed in ORS Dragonfly software. The measured distances between the cochlear boundary and specimen edges were aligned with the intended safety margin of 5 mm, with an average error of ±0.85 mm. Expert review of the overall anatomy revealed no evidence of structural damage. All cochleae and semicircular canals were intact, as were any implants. Our process streamlined prior methods, sectioning bone samples up to 3 cm wide in under 3 minutes. Utilizing a laser cutter expedites sample preparation, extracting uniform sections and preserving delicate internal structures, while minimizing risk of damage. This method accelerates high-resolution imaging workflows and improves the accuracy of structural analysis in diverse clinical specimens.

Atherosclerosis is the primary underlying cause of cardiovascular disease and exhibits clear sex differences in onset and severity. While gonadal hormones contribute to these differences, sex chromosomes may independently influence risk. Kdm5d encodes a Y-linked histone demethylase that is highly expressed in plaques, yet its functional role in disease progression remains undefined. To test whether Kdm5d influences atherosclerosis, we generated C57BL6/J Apoe-/- mice overexpressing Kdm5d (Kdm5d-Tg) and compared them to Apoe-/- littermates in both sexes. We measured body weight, body composition, plasma lipids, glucose, circulating immune cells, and aortic sinus lesion size. Kdm5d overexpression changed body weight in both sexes without altering body composition. In females, Kdm5d significantly reduced plasma triglyceride and cholesterol levels, whereas in males, these metabolic measures were unchanged. In contrast, male Kdm5d-Tg mice showed increased immune cell counts, including white blood cells and monocytes, with no such change in females. Kdm5d overexpression did not significantly alter atherosclerotic lesion size in either sex. Together, these findings show sex-specific metabolic and immune effects of Kdm5d and implicate Y chromosome gene function as a gonadol hormone-independent contributor to sex differences in atherosclerosis. As this is a global overexpression model, future studies using cell-type-specific approaches will be necessary to dissect which Kdm5d-expressing populations drive the observed phenotypes.

The tumor suppressor p53 is a central regulator of cell cycle progression and genomic stability, primarily through transcriptional activation of p21, a cyclin dependent kinase inhibitor. In pulmonary hypertension, dysregulated cell proliferation contributes to vascular remodeling, in which fibroblasts play a key role in pathological cell growth. As a downstream effector of p53, p21 inhibits cyclin CDK complexes to regulate cell cycle arrest under stress conditions. This study investigates the role of p21 in fibroblast cell cycle regulation and its contribution to p53 signaling in the context of pulmonary hypertension. To assess p21 function, small interfering RNA was used to knock down p21 expression in cultured fibroblasts. Western blot analysis was then performed to evaluate changes in p21 protein levels and downstream cell cycle regulators, allowing assessment of pathway activity following gene silencing. We hypothesize that p21 knockdown will reduce inhibition of cyclin CDK complexes, resulting in increased cell cycle progression. These findings will further define the role of p21 as a mediator of p53 dependent signaling in fibroblast proliferation. This work provides insight into mechanisms of vascular remodeling in pulmonary hypertension and may inform future therapeutic strategies targeting the p53 p21 pathway.