Date of Award

12-2025

Degree Type

Honors College Thesis

Academic Program

Biological Sciences BS

Department

Biological Sciences

First Advisor

Ramesh Rijal

Advisor Department

Biological Sciences

Abstract

Tuberculosis (TB) kills 1.5 million people annually, with males experiencing higher disease incidence and worse clinical outcomes than females. Despite this disparity, the cellular and molecular mechanisms underlying sex-specific differences in TB immunity remain poorly understood. Our lab previously observed that polyphosphate (polyP), a polymer of phosphate secreted by pathogens such as Mycobacterium tuberculosis (Mtb), enhances the survival of ingested bacteria in human macrophages and alters macrophage gene expression in a sex-specific manner. This study investigated how Mtb infection or polyP treatment of macrophages alters the expression of Ceroid lipofuscinosis, neuronal 8 (CLN8), one of the most highly altered genes identified in our previous gene expression analysis. CLN8 encodes an endoplasmic reticulum-localized lipid trafficking protein, suggesting a potential role in lipid remodeling and stress responses during infection. We therefore focused on CLN8 to explore its contribution to lipid droplet accumulation and nuclear morphology in macrophages.

Using confocal microscopy, we analyzed macrophages from three male and three female donors treated with control, polyP, or Mtb. Contrary to our previous RNA sequencing data showing sex-specific CLN8 transcriptional upregulation in females, CLN8 protein expression showed no significant changes in either sex across treatments. However, clear sex-specific differences in phenotypic lipid metabolism emerged. Both male and female macrophages accumulated lipid droplets in response to polyP treatment, but only females showed significant lipid accumulation during Mtb infection, indicating impaired signal integration in males. Nuclear morphology analysis showed that female macrophages possess significantly rounder nuclei than males at baseline—a stable architectural difference maintained across all treatments. Males uniquely showed decreased nuclear roundness following polyP treatment, suggesting sex-specific structural responses to bacterial signals.

These findings demonstrate that biological sex fundamentally shapes macrophage responses to TB through distinct cellular strategies: females exhibit robust metabolic flexibility with a stable nuclear architecture, while males show threshold-dependent responses that require strong, isolated signals. The identification of polyP as a key mediator of sex-specific cellular responses provides mechanistic insight into clinical TB disparities and suggests that personalized, sex-specific treatment approaches may improve outcomes for this deadly disease.

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