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. 2018 Dec 11:9:2881.
doi: 10.3389/fimmu.2018.02881. eCollection 2018.

The Therapeutic Effect of STAT3 Signaling-Suppressed MSC on Pain and Articular Cartilage Damage in a Rat Model of Monosodium Iodoacetate-Induced Osteoarthritis

Seon-Yeong Lee  1 , Seung Hoon Lee  1 , Hyun Sik Na  1 , Ji Ye Kwon  1 , Goo-Young Kim  1 , KyungAh Jung  2 , Keun-Hyung Cho  1 , Seon Ae Kim  3 , Eun Jeong Go  3 , Min-Jung Park  1 , Jin-Ah Baek  1 , Si Young Choi  1 , JooYeon Jhun  1 , Sung-Hwan Park  1   4 , Seok Jung Kim  3 , Mi-La Cho  1   2   5   6
Affiliations

The Therapeutic Effect of STAT3 Signaling-Suppressed MSC on Pain and Articular Cartilage Damage in a Rat Model of Monosodium Iodoacetate-Induced Osteoarthritis

Seon-Yeong Lee et al. Front Immunol. .

Abstract

Osteoarthritis (OA) is a degenerative disease that induces pain, cartilage deformation, and joint inflammation. Mesenchymal stem cells (MSCs) are potential therapeutic agents for treatment of OA. However, MSC therapy can cause excessive inflammation. Signal transducer and activator of transcription 3 (STAT3) modulates secretion of many proinflammatory cytokines. Experimental OA was induced by intra-articular (IA) injection of monosodium iodoacetate (MIA) to the right knee of rats. MSCs from OA patients (OA-MSCs) were treated with STA21, a small molecule that blocks STAT3 signaling, by IA or intravenous (IV) injection after MIA injection. Pain severity was quantified by assessment of secondary tactile allodynia using the von Frey assessment test. Cartilage degradation was measured by microcomputed tomography image analysis, histological analysis, and the Mankin score. Protein and gene expression was evaluated by enzyme-linked immunosorbent assay, immunohistochemistry, and real-time polymerase chain reaction. MSCs increased production of proinflammatory cytokines under inflammatory conditions. STA21 significantly decreased expression of these proinflammatory molecules via inhibition of STAT3 activity but increased gene expression of molecules related to migration potential and immunomodulation in OA-MSCs. STAT3-inhibited OA-MSCs administrated by IV or IA injection decreased pain severity and cartilage damage in rats with MIA-induced OA rats by decreasing proinflammatory cytokines in the joints. Combined IA and IV-injected STAT3-inhibited OA-MSCs had an additive effect of pain relief in MIA-induced OA rats. STAT3 inhibition may optimize the therapeutic activities of MSCs for treating OA by attenuating pain and progression of MIA by inhibiting inflammation and cartilage damage.

Keywords: TRPV1; inflammation; mesenchymal stem cells; osteoarthritis; stat3.

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Figures

Figure 1
Figure 1
Effects of STAT3 inhibition on anti-inflammatory activity and migration potential in MSCs from osteoarthritis (OA) patients. (A) OA-MSCs were incubated with STA21 for 3 days and, ELISA was performed to measure IL-6, IL-8, IL-1β, and VEGF concentrations. (B) OA-MSCs were incubated with STA21 for 1 h. The expression of phospho (p)-STAT3 705, 727, and total STAT3 protein was analyzed by Western blotting. (C) mRNA levels for anti-inflammatory cytokines IL-10 and TGF-β were measured by real time-PCR (D) Expression of chemokine receptors in OA-MSCs incubated with STA21 was measured by real time-PCR. OA-MSC were isolated of fat tissues obtain from each 3 person with OA. Data represent the mean ± SD of 3 independent experiments (*P < 0.05; **p < 0.01).
Figure 2
Figure 2
Reduced severity by IA administration of iSTAT3 OA MSCs in MIA-induced OA rats. (A) OA was induced in Wistar rats by IA injection of MIA. OA Rats were injected with IA of normal (Nor-) MSCs (6 × 105), OA-MSCs (6 × 105), or iSTAT3 OA-MSCs (6 × 105). Nor-MSC were isolated of fat tissues obtain from each 2 normal person, and OA-MSC were from each 4 person with OA. Pain behavior was assessed as mechanical hyperalgesia measured with a dynamic plantar esthesiometer and incapacitance meter, and was quantified as PWL and PWT (n = 3 per group). (B) Representative micro-CT images of the femoral condyles at 4 weeks in all groups. Object volume (Obj.V) and bone surface (%) were measured in micro-CT images from femurs. (*P < 0.05, **P < 0.01, ***P < 0.001). (C) Histochemical analysis of Femoral condyle and severity scores after IA administration of iSTAT3-OA MSCs to MIA-induced OA rats. Knee joint tissue samples were acquired from all OA groups at 4 weeks and stained with H&E and Safranin O to evaluate the severity of inflammation and cartilage damage (*P < 0.05, **P < 0.01, ***P < 0.001).
Figure 3
Figure 3
Reduced severity by IV administration of iSTAT3 OA-MSCs in MIA-induced OA rats. (A) OA was induced in Wistar rats by IA injection of MIA. OA rats were injected IV with OA-MSCs (6 × 105) or iSTAT3 OA-MSC (6 × 105). OA-MSC were isolated of fat tissues obtain from each 3 person with OA. Pain behavior was assessed as mechanical hyperalgesia measured with a dynamic plantar esthesiometer and incapacitance meter, and was quantified as PWL and PWT (n = 3 per group). (B) Representative micro-CT images of the femoral condyles at 4 weeks in all groups. Object volume (Obj.V) and bone surface (%) measured in micro-CT images from femurs. (C) Histochemical analysis of femoral condyle and severity scores after IV administration of iSTAT3-OA MSCs in MIA-induced OA rats. Knee joint tissue samples were acquired at 4 weeks from wild-type (WT) rats, rats with MIA-induced OA, rats that received IV OA-MSCs, and rats that received IV iSTAT3 OA-MSCs, and were stained with H&E and Safranin O to evaluate the severity of inflammation and cartilage damage (*P < 0.05; **P < 0.01; ***P < 0.001).
Figure 4
Figure 4
Additive effects of pain relief by IA and IV co-administration of iSTAT3 OA-MSCs in MIA-induced OA rats. (A) OA was induced in Wistar rats by IA injection of MIA. OA rats were injected with IA and IV iSTAT3 OA-MSCs (each 3 × 105, final 6 × 105 per rat). OA-MSC were isolated of fat tissues obtain from each four person with OA. Pain behavior was assessed as mechanical hyperalgesia measured with a dynamic plantar esthesiometer and incapacitance meter, and was quantified as PWL and PWT (n = 3 per group). (B) Immunohistochemical staining was used to identify the expression of TRPV1 in the dorsal root ganglion of OA rats. *P < 0.05, **P < 0.01, ***P < 0.001.
Figure 5
Figure 5
Additive effect of IA and IV co-administration of iSTAT3 OA-MSCs on joint protection in MIA-induced OA rats. (A) Representative micro-CT images of the femoral condyles at 4 weeks in all groups. Object volume (Obj.V) and bone surface (%) from micro-CT images of the femur (*P < 0.05, ***P < 0.001). (B,C) Histochemical analysis of femoral condyle (B) and severity scores (C) after IA and IV co-administration of iSTAT3-OA MSCs in MIA-induced OA rats. (B) OA was induced in Wistar rats by IA injection of MIA. OA Rats were injected multiple times with IA and IV co-administration of iSTAT3 OA-MSCs (3 × 105). Knee joint tissue samples were acquired at 4 weeks from wild-type (WT) rats, rats with MIA-induced OA, rats that received IV OA-MSCs, and rats that received IV iSTAT3 OA-MSCs, and were stained with H&E and Safranin O to evaluate the severity of inflammation and cartilage damage (***P < 0.001).
Figure 6
Figure 6
Expression of inflammatory mediators by administration of STAT3-inhibited OA-MSCs in articular cartilage of MIA-induced OA rats. Immunohistochemical staining of femoral condyle was performed to measure the expression of IL-1β, IL-6, TNF-α, and HIF-2α after multiple injections of IA and IV iSTAT3 OA-MSCs in knee joint tissues of MIA-induced OA rats. The percentage positive cells for each antibody are shown at the right. Data represent the mean ± SEM of 3 independent experiments (***P < 0.001).
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