Chondrogenic potential of mesenchymal stem cells from horses using a magnetic 3D cell culture system

BACKGROUND Mesenchymal stem cells(MSCs)represent a promising therapy for the treatment of equine joint diseases,studied due to their possible immunomodulatory characteristics and regenerative capacity.However,the source of most suitable MSCs for producing cartilage for regenerative processes in conjunction with biomaterials for an enhanced function is yet to be established.AIM To compare the chondrogenicity of MSCs derived from synovial fluid,bone marrow,and adipose tissue of horses,using the aggrecan synthesis.METHODS MSCs from ten horses were cultured,phenotypic characterization was done with antibodies CD90,CD44 and CD34 and were differentiated into chondrocytes.The 3D cell culture system in which biocompatible nanoparticles consisting of gold,iron oxide,and poly-L-lysine were added to the cells,and they were forced by magnets to form one microspheroid.The microspheroids were exposed to a commercial culture medium for 4 d,7 d,14 d,and 21 d.Proteoglycan extraction was performed,and aggrecan was quantified by enzyme-linked immunosorbent assay.Keratan sulfate and aggrecan in the microspheroids were identified and localized by immunofluorescence.RESULTS All cultured cells showed fibroblast-like appearance,the ability to adhere to the plastic surface,and were positive for CD44 and CD90,thus confirming the characteristics and morphology of MSCs.The soluble protein concentrations were higher in the microspheroids derived from adipose tissue.The aggrecan concentration and the ratio of aggrecan to soluble proteins were higher in microspheroids derived from synovial fluid than in those derived from bone marrow,thereby showing chondrogenic superiority.Microspheroids from all sources expressed aggrecan and keratan sulfate when observed using confocal immunofluorescence microscopy.All sources of MSCs can synthesize aggrecan,however,MSCs from synovial fluid and adipose tissue have demonstrated better biocompatibility in a 3D environment,thus suggesting chondrogenic superiority.CONCLUSION All sources of MSCs produce hyaline cartilage;however,the use of synovial liquid or adipose tissue should be recommended when it is intended for use with biomaterials or scaffolds.

The role of corneal endothelium in macular corneal dystrophy development and recurrence

Macular corneal dystrophy(MCD)is a progressive,bilateral stromal dystrophic disease that arises from mutations in carbohydrate sulfotransferase 6(CHST6).Corneal transplantation is the ultimate therapeutic solution for MCD patients.Unfortunately,postoperative recurrence remains a significant challenge.We conducted a retrospective review of a clinical cohort comprising 102 MCD patients with 124 eyes that underwent either penetrating keratoplasty(PKP)or deep anterior lamellar keratoplasty(DALK).Our results revealed that the recurrence rate was nearly three times higher in the DALK group(39.13%,9/23 eyes)compared with the PKP group(10.89%,11/101 eyes),suggesting that surgical replacement of the corneal endothelium for treating MCD is advisable to prevent postoperative recurrence.Our experimental data confirmed the robust m RNA and protein expression of CHST6 in human corneal endothelium and the rodent homolog CHST5 in mouse endothelium.Selective knockdown of wild-type Chst5 in mouse corneal endothelium(AC^(siChst5)),but not in the corneal stroma,induced experimental MCD with similar extracellular matrix synthesis impairments and corneal thinning as observed in MCD patients.Mice carrying Chst5 point mutation also recapitulated clinical phenotypes of MCD,along with corneal endothelial abnormalities.Intracameral injection of wild-type Chst5 rescued the corneal impairments in AC^(siChst5)mice and retarded the disease progression in Chst5 mutant mice.Overall,our study provides new mechanistic insights and therapeutic approaches for MCD treatment by highlighting the role of corneal endothelium in MCD development.