CRYOPRESERVATION OF ENDOMETRIAL STROMAL CELLS OF BUFFALO (BUBALUS BUBALIS)

The aim of the present experiment was to cryopreserve buffalo endometrial stromal cells and to study their morphological and physiological characteristics. Stromal cells isolated by enzymatic digestion from the buffalo uterus of day 1-5 of the estrous cycle, were cultured and frozen at -80 degrees C using DMSO as a cryoprotectant. Frozen cells were thawed at 37 degrees C and recultured. The culture media was collected when both unfrozen (control) and frozen cells reached confluence for estimation of PGE(2) and PGF(2 alpha) using enzyme-linked immunosorbent assay. Frozen stored buffalo endometrial stromal cells immediately after thawing appeared as single or in small clumps, glistening and rounded which changed their morphology from rounded to flat spindle shaped immediately 24 h after seeding retaining their normal morphological characteristics. The viability of frozen thawed stromal cells at the time of seeding estimated by Trypan blue dye exclusion was 63.3%. There was no significant difference in the basal production of PGE(2) and PGF(2 alpha) by the frozen-thawed and unfrozen buffalo stromal cells (289.38 +/- 4.39, 3.28 mu 0.16 pg/mu g protein vs. 305.97 +/- 3.20, 4.26 +/- 0.17 pg/mu g protein, respectively) on day 7 of culture indicating no adverse effect of cryopreservation on their functional characteristics. The results demonstrated that buffalo endometrial stromal cells can be cryopreserved without any major impairment in their morphological and functional integrity.