Biochemistry

2509 Submissions

[2] viXra:2509.0157 [pdf] submitted on 2025-09-30 23:25:15

A Method [for Mapping] Cell Lineage with Mutant Mitochondrial Transfer

Authors: Jaba Tkemaladze
Comments: 15 Pages. (Note by viXra Admin: Please don't name the method after the author's name - Please conform))

The comprehensive mapping of cellular lineages from the zygote to a fully formed organism remains a fundamental and unresolved challenge in developmental biology. While modern single-cell technologies offer snapshots of cellular heterogeneity, they lack the inherent, permanent markers required to trace progeny through the complex events of asymmetric division and migration over time. This work introduces the Tkemaladze Method, a novel lineage-tracing approach that utilizes mutant mitochondrial DNA (mtDNA) as a stable, inheritable genetic label. The method involves the isolation of mitochondria from cytoplasts harboring known pathogenic mtDNA mutations and their microinjection into murine embryonic stem cells (mESCs). We confirmed successful transfer and functional integration via fluorescence microscopy and quantitative PCR. These labeled progenitor cells were used to generate chimeric embryos, where we demonstrated stable heteroplasmy and faithful inheritance of the mutant mtDNA in clonal progeny throughout development. Using fluorescent reporters, we visualized the fate of individual progenitors, enabling the quantitative construction of a detailed cytogenealogical map across tissues like the central nervous system, liver, and myocardium. A key finding was the tissue-specific segregation of mitochondrial tags, revealing selective pressure in high-energy-demand tissues. The Tkemaladze Method thus provides an unprecedented, powerful tool for fundamental developmental biology, disease modeling, and tracking the fate of transplanted cells in regenerative medicine.
Category: Biochemistry

[1] viXra:2509.0088 [pdf] submitted on 2025-09-15 19:51:12

Production of Functional Gametes from Somatic Cells of the Planarian Schmidtea Mediterranea Via in Vitro Gametogenesis

Authors: Jaba Tkemaladze, Gabro Gakely, Laura Gegelia, Iason Papadopulo, Alexander Taktakidze, Nino Metreveli, Natia Berozashvili, Natalia Bondarenko, Ucha Maglakelidze
Comments: 25 Pages.

This study establishes a novel and robust protocol for the direct reprogramming of differentiated somatic cells into functional gamete precursors in the planarian Schmidtea mediterranea, bypassing the need for a pluripotent intermediate state. Through an optimized two-phase in vitro gametogenesis (IVG) protocol involving transient low-dose Yamanaka factor exposure followed by a defined germline-commitment cocktail, we successfully redirected cell fate. Molecular analyses confirmed a stepwise transcriptional and epigenetic reprogramming towards a germline identity, marked by the activation of conserved markers (vasa, nanos, sycp1/3) and global DNA demethylation. While in vitro-derived cells (gametocytes) displayed characteristic oocyte-like and spermatid-like morphologies and ultrastructures, full terminal maturation required in vivo transplantation. Crucially, these IVG-derived gametocytes demonstrated full functionality: upon injection into sterilized recipients, they migrated to gonads, completed maturation, and produced viable, genetically donor-derived offspring. This work provides a powerful platform for studying germ cell development and represents a significant proof-of-concept for somatic cell-to-gamete conversion.
Category: Biochemistry