An Enriched Primary Culture Reveals Morphological Heterogeneity of Rat Testicular Telocytes.

Telocytes are stromal cells characterized by small cell bodies and long cytoplasmic extensions termed telopodes. Although identified in the rat testicular interstitium, no standardized protocol for their isolation and in vitro enrichment has been established. This study aimed to develop an enriched primary culture model for rat testicular telocytes and characterize their morphology in vitro. Testicular cells from adult male Wistar rats were isolated by enzymatic digestion, subjected to short-term culture with differential adhesion, and enriched by fluorescence-activated cell sorting based on a CD34-positive/CD31-negative profile with empirical exclusion of events within the αSMA-associated region to reduce PTMC-like contamination. Post-sorting CD34 immunoreactivity, absence of detectable CD31 immunoreactivity, and characteristic morphology supported interpretation of the sorted fraction as telocyte-enriched. Light microscopy revealed recurring stellate, bipolar, compact branched, chain-like, and dense network-forming patterns. Some long, thin cytoplasmic processes showed a moniliform appearance with alternating thin segments and focal dilations. The findings demonstrate culture-associated morphological heterogeneity within the enriched population and provide a methodological framework for its isolation and light-microscopic characterization. Expanded post-sorting phenotyping, including PDGFRα and vimentin, together with ultrastructural validation, will be required in future studies.