Hyaluronan synthase-2-specific antisense oligonucleotides inhibit the formation of extracellular coat in human peritoneal mesothclial cells / 中华肾脏病杂志

ABSTRACT

Objective To define which hyaluronan synthase (HAS), of three hyaluronan synthesizing enzymes HAS-1, HAS-2, and HAS-3, is primarily responsible for hyaluronan synthesis and extracellular matrix/extracellular coat formation in human peritoneal mesothelial cells (HPMCs) . Methods As a prerequisite study, the expression of each HAS mRNA in cultured HPMCs was measured by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR). Only the expression of HAS-2 and HAS-3 mRNA could be detected. The level of HAS-2 mRNA expression was about 10 fold higher than that of HAS-3. HAS-2 specific antisense oligonucleotide was then transfected into cultured HPMCs by lipofectamine. After 0, 8, 24, and 48 hours, the expression of HAS-2 mRNA was detected by RT-PCR and the extracellular coat was measured by particle exclusion test. Results 8 hours and 24 hours after transfection, the expression of HAS-2 mRNA in HPMCs decreased by 58% and 89% respectively; 48 hours after transfection, the expression of HAS-2 mRNA in HPMCs partially restored to 25% of the normal level. Correspondingly, 24 hours after transfection, the extracellular matrix/extracellular coat in HPMCs almost completely disappeared. However, as control, sense and reverse oligonucleotides showed no effect. Conclusion HAS-2 plays a leading role in HPMCs hyaluronan synthesis and the formation of extracellualr matrix/extracellular coat.