Background/Purpose: Hexokinases (HKs) catalyze the first step in glucose metabolism. HK2 constitutes the principal inducible isoform with a restricted distribution in normal adult tissues. Up-regulated glycolysis and HK2 expression have a powerful growth advantage, which promotes cell proliferation and invasion. Fibroblast-like synoviocytes (FLS) are a key component of rheumatoid arthritis (RA) invasive synovium, and display unique aggressive features, including increased migration and invasion. We have recently showed that RA FLS present an altered glucose metabolism, which modulates FLS mediated functions. To determine whether HK2 is a key regulator of FLS glycolytic and aggressive phenotype, we examined its role on FLS functions and in an arthritis animal model.

Methods: HK1 and HK2 expression were studied by immunohistochemistry (IHC) in RA and osteoarthritis (OA) synovial tissue. HK2 expression in FLS was studied by double IHC and qPCR in synovial tissue and FLS cell lines, respectively. RA FLS were transfected with HK2 siRNA, or infected with HK2 expressing adenovirus (ad) and the following FLS functions were conducted: migration (scratch assay) and matrigel invasion assay after PDGF stimulation, and metalloproteases (MMP) expression after LPS stimulation. We used HK2^F/Fmice, harboring the ubiquitously expressed tamoxifen inducible Cre (UBCcreERT2) in order to systemically delete HK2. Passive KRN arthritis was induced injecting intraperitoneally 150μl of serum 3 weeks after tamoxifen treatment. Clinical scores were assessed daily and histopathological studies were carried on day 10.

Results:  While HK1 is widely expressed in both RA and OA synovial tissue, HK2 is particular of RA histopathology (9/9 RA; 3/9 OA). HK2 is localized in the synovial lining and sublining and it co-localizes with vimentin as a marker for FLS. Basal HK2 expression was increased in RA compared to OA FLS, and its expression increased after LPS and PDGF stimulation. Silencing HK2 in RA FLS resulted in a less invasive phenotype (3.2 fold less invaded area in HK2-silenced FLS, p<0.05) and lower levels of MMP3 expression. Consistently, overexpression of HK2 resulted in an increased ability to migrate (scratch length: GFP-ad: 299.1±7.2; HK2-ad 90.7±2.5; p<0.001) and invade (3.1 fold invaded area in HK2-ad; p<0.001). The UBCcre-HK2^F/F mice showed lower clinical and histological arthritis scores than wild type mice (WT) (clinical score at day 10: WT: 9±0.79; UBCCre-HK2^F/F: 3±0.49 (p<0.001)).

Conclusion: HK2 is more abundantly expressed in RA than OA synovial lining, and regulates FLS aggressive functions. HK2 also contributes to arthritis severity in the KRN animal model. Therefore, selective HK2 inhibition might be an attractive potential selective target safer than global glycolysis.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/hexokinase-2-as-a-novel-metabolic-target-for-rheumatoid-arthritis/