Nowadays, all guidelines agree that non-pharmacological treatments such as education and self-management, exercise, weight control, and walking aids should be central to the management of patients with OA (Zhang et al., 2010; Nelson et al., 2014). Most of the guidelines recommend strongly that OA patients should acquire up-to-date information and education to allow them to self-manage the disease to a certain extent (Zhang et al., 2008; Block, 2014). Exercise therapy (strengthening exercise and aerobic exercise) is helpful in reducing pain, improving joint motion, and strengthening muscles around the joints (Jordan et al., 2003). Obesity or overweight is associated with the prevalence and progression of knee OA, while weight loss can help to relieve their OA symptoms and delay the structural damage (Messier, 2009). The benefits of knee braces and other assistive devices for physical support and assistance are still controversial and not well-organized (Thomas et al., 2018). Besides, some alternative medicine treatments, like acupuncture, thermal modalities, and therapeutic ultrasound are likely to have little effect in reducing the pain of OA patients (Block, 2014).

Clinical evidence also suggests that some of OA patients will benefit from drugs (Ringdahl and Pandit, 2011). Drugs, including acetaminophen, non-steroidal anti-inflammatory drugs (NSAIDs), and opioids are essential medicine for patients who have moderate to severe pain. Among these drugs, acetaminophen and NSAIDs are recommended as the first-line pain medication for OA by most guidelines (Dougados, 2006; Hunter and Bierma-Zeinstra, 2019). Nonetheless, safety should be an important consideration in selecting these drugs, since they are reported to be related to considerable side effects, such as liver toxicity, gastrointestinal and cardiovascular complications (Sostres et al., 2010). Opioids are more potent and effective drugs for patients with refractory pain. Both short and long-acting opiates are effective in managing OA pain and have level 3 evidence in their support (Ringdahl and Pandit, 2011). Benefits from the opiates may be acquired, however, frequent adverse effects are associated with these drugs including nausea, dizziness, vomiting, constipation, and sleepiness (Fuggle et al., 2019). In addition, concerns about pharmacologic tolerance, physical dependence suggest the use of opioids should be appropriately dosed and monitored (Lipman, 2001).

There is emerging evidence that IA injections of CSs and HA are helpful for some OA patients (Wernecke et al., 2015; Concoff et al., 2017). CS is known to inhibit the release of inflammatory cytokines in the affected joint and restrain further cartilaginous destruction (Song et al., 2012). IA injections of CSs may provide some patients with temporary symptomatic relief, and a low risk of adverse effects (Wernecke et al., 2015). According to the guideline from Osteoarthritis Research Society International (OARSI), CSs injections should be performed after patients failing or are unsatisfactory with oral analgesic/anti-inflammatory agents, especially for the patients with symptomatic knee OA with effusions or other physical signs of local inflammation (Zhang et al., 2008). HA is a constituent of synovial fluid, while the contents of HA are decreased and compromised in OA joints (Temple-Wong et al., 2016; Chen et al., 2020). Exogenous supplementation of HA is thought to be a visco-supplemental or pharmaceutical therapy for patients with knee OA. They are inferior to CSs in terms of short-term duration, but with likely prolonged symptomatic benefit (Trueba Davalillo et al., 2015). Many complementary medicines (glucosamine sulphate, chondroitin sulphate, ginger, turmeric, etc.) and nutritional supplements have been used to treat OA, but little detail was given and there was no consensus has been achieved (Nelson et al., 2014).

The efficacy of MSC-based therapy has been proven to be effective in different OA animal models (Gupta et al., 2012; Grassel and Lorenz, 2014; Sasaki et al., 2019), ranging from murine, rabbit, canine, ovine, and caprine to equine (see Table 2). In vivo study, IA injection of human adipose tissue-derived MSCs (AD-MSCs) engrafted into the rat joints and increased the cartilage thickness in surgery-induced OA animal models (Li et al., 2016). Small animal models are often used as a proof-of-concept, owing to their AC is thinner and smaller than humans. Large animals, such as ovine, equine, or sheep are more suitable for modeling human AC defects and investigating the effectiveness of MSC-based treatments (Grassel and Lorenz, 2014). In a study conducted by Al Faqehand colleagues, autologous bone marrow MSCs (BM-MSCs) were performed for IA injection of surgically-induced OA sheep. 6 weeks after injection, gross evidence of retardation of cartilage destruction was observed in the OA joints treated with BM-MSCs (Al Faqeh et al., 2012). Consistent with this study, the study of Ko et al. found that injected SOX-6, 9-transfected AD-MSCs attenuated the progression of OA in goats (Ko et al., 2019). The safety and feasibility of IA injection of MSCs for cartilage repair have also been evaluated in several clinical trials (Grassel and Lorenz, 2014; Lee and Wang, 2017). It is suggested that subjects with mild to moderate OA (K-L grade 1–3) are optimal candidates for MSC therapy (Lee and Wang, 2017). Jo et al. enrolled 18 patients with knee OA (K-L grade 2–3) and injected autologous AD-MSCs into the knee, showing improved Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) score, decreased size of the cartilage defect, and increased cartilage volume at 6 months after injection in the high-dose group (1.0 × 10⁸/L) (Jo et al., 2014). Concomitantly, in a 2 years’ follow-up study, Orozco et al. found that IA injection of autologous expanded BM-MSCs was effective in improving both magnetic resonance imaging (MRI) T2 mapping and visual analogue scale (VAS) outcomes in patients undergoing chronic knee pain (Orozco et al., 2013; 2014). In addition, one randomized controlled trial (RCT) study assessing the feasibility and safety of treating OA with allogeneic BM-MSCs was reported by (Vega et al. (2015) . In this study, 30 patients with chronic knee OA were randomized into 2 groups of 15 patients: the test group received allogeneic BM-MSCs (40 × 10⁶ cells) by IA injection, while the control group received HA (60 mg). After 12 months, the test group illustrated significant improvement in algofunctional indices and quantification of cartilage quality compared to the control group (Vega et al., 2015). Nevertheless, patients withstanding larger cartilage lesions exhibited significantly inferior consequences (Im, 2018). Additionally, MSCs obtained from subjects with severe OA have decreased potential for proliferation and chondrogenic differentiation, resulting in a lower cell yield and higher osteogenic differentiation (Murphy et al., 2002).

Although encouraging symptomatic relief has been demonstrated in clinical reports, hyaline AC regeneration was rarely reported. In a study with three case reports, Wakitani et al. found that the cartilage defect had been repaired with the fibrocartilaginous tissue in the first patient after 1 year of BM-MSCs transplantation. MRI results of the second patient revealed complete coverage of the defect, while not able to determine the covered materials was hyaline cartilage (Wakitani et al., 2007). Discoveries from animal researches implicated that MSCs achieve therapeutic effects in virtue of paracrine mechanisms, rather than integrating with cartilage directly and producing ECM in situ (Saulnier et al., 2015; Ozeki et al., 2016). These paracrine factors consist of various proteins and could be conveyed in extracellular vehicles (EVs) (Lee and Wang, 2017). There are three major categories of EVs: apoptotic bodies, exosomes or nanovesicles, and microparticles/microvesicles (MPs) (Mianehsaz et al., 2019). Recently, it has been increasingly supported that MSCs-derived exosomes contribute to the reparative effects of MSC-based treatment in OA models (Cosenza et al., 2017) (Zhang et al., 2019). An in vivo study revealed that BM-MSCs and MSCs-derived exosomes equally protected collagenase-induced mice from joint damage (Cosenza et al., 2017). Other issues such as frequency of injection, the dose of MSCs, MSC origin, and transplantation type also should be taken into consideration (Im, 2018). Moreover, the safety of IA injection of MSCs is one of the key points in clinical application. In one meta-analysis, Peeters assessed the reported adverse events of IA treatment with cultured stem cells in humans, four serious adverse effect cases were reported, including one infection, one pulmonary embolism, and two tumors (Peeters et al., 2013). Lastly, a lacking of a large number of multicenter data and high-level evidence hinders the application of MSCs in early-stage OA. Given current knowledge, the preliminary results indicated that IA MSCs injection is promising in reducing pain and improving the quality of OA patients. However, more RCTs and high-level evidence are still required before in-depth clinical translation.

Notably, over the past decade, the cartilage regeneration potential of hydrogels combined with or without cells and biologics have been investigated and remarkable successes have been achieved in fundamental studies (Roberts et al., 2011; Kontturi et al., 2014; Arora et al., 2017). Kontturi et al. designed an injectable, in situ forming type II collagen/HA hydrogel and found that it could assist the long-term survival of the encapsulated chondrocytes as well as maintain their round shape in vitro (Kontturi et al., 2014). In another study, Roberts et al. demonstrated that a chondrocyte-laden PEG-LA hydrogel (consisting of PEG and oligomers of lactic acid (LA)) significantly improved the formation of cartilage-like tissue comprised of glycosaminoglycan and collagen under loading (Roberts et al., 2011). Moreover, Arora et al. have reported that TGF-β1 loaded hydrogels enhanced cell survival and chondrogenesis of MSCs and chondrocytes (Arora et al., 2017). In parallel, many preclinical studies have assessed the reparative effects of injectable hydrogels on the cartilage in large and small animal models (Na et al., 2007; Wang et al., 2007; Vilela et al., 2015). Na et al. developed a thermo-reversible hydrogel construct blended with HA, which was used as an injectable carrier for rabbit chondrocytes and TGF-β3. The results showed that when the blended hydrogels mixed with TGF-β3 and chondrocytes were injected subcutaneously into the nude mice, the level of cartilage associated ECM production was significantly higher than those without HA or TGF-β3 (Na et al., 2007). Wang et al. used a chondroitin sulphate (CS) based adhesive-hydrogel to stimulate cartilage formation in goat models. This study suggested greater defects fill in the presence of the hydrogels compared to microfracture alone (Wang et al., 2007).

Clinical trials of injectable hydrogels on OA treatment are very limited, and their applications are mainly to boost the efficacy of existing therapies, such as microfracture, osteochondral grafting (Wu et al., 2020). Most of the reported trials are case reports or case series. In a pilot clinical study, Sharma et al. recruited 18 subjects with focal cartilage defects on the medial femoral condyle. 15 subjects (treated group) were treated with the adhesive-hydrogel in conjunction with microfracture, while three subjects (control group) were treated with microfracture alone. At 6 months follow up, MRI analysis showed significant improvement of repair tissue integration was achieved in the treated group. The treated group also experienced pain reduction compared to the controls, and no major adverse events were observed (Sharma et al., 2013). Another clinical trial using ChonDux hydrogel (consisting of a PEG/HA network and a CS adhesive) for treating full-thickness femoral condyle defects (Wolf et al., 2020). Researchers found that ChonDux mediated 94.2 ± 16.3% of final defects fill over 2-years of follow-up and the treated tissue was similar to adjacent cartilage between 12 and 24 months, suggesting ChonDux is a safe accessory to microfracture therapy (Wolf et al., 2020). In addition, a novel medicine product (Cartistem), comprised of HA hydrogel and allogeneic human umbilical cord blood-derived MSCs (hUCB-MSCs) was assessed in a clinical study over 7-years of follow-up. The results suggested this product appears to be safe and effective for cartilage regeneration in knee OA (Park et al., 2017). The above pilot studies confirmed the prospects of injectable hydrogel in the treatment of OA. However, some challenges for clinical translation remain to be addressed, including the source of the material, and integration to the cartilage in a stable way. Future studies are needed to address these issues before clinical application.

Delivery of the transgene can be achieved by viral or non-viral methods. At present, viral vectors are more preferred and have been used for IA gene delivery in animal models (Kay et al., 2009; Wang et al., 2016) and human trials (Mease et al., 2009; Mease et al., 2010). Retroviruses and adeno-associated virus (AVV) are the only vectors that have been assayed in clinical studies until now. Compared with other viral vectors, AAV has the advantage of penetrating deeply within cartilage to transduce chondrocytes in situ (Madry et al., 2003). In vivo experiments by Kay et al. showed that the level of IL-1Ra expression present in the joint space of self-complementary AAV (sc-AAV) injected rabbits for 2 weeks in sufficient quantities to suppress inflammation of the IL-1β-induced arthritis model (Kay et al., 2009). Similarly, in another study, Wang et al. suggested that no local or systemic toxicity was found following sc-AAV vector carrying IL-1Ra transgene (sc-rAAV2.5IL-1Ra) injection in mono-iodoacetate (MIA)-induced OA rats (Wang et al., 2016). Sustained expression of IL-1Ra and a low rate of cartilage loss were observed in the vector-injected knees (Wang et al., 2016). In large mammalian joints, transgenic expression of human IL-1Ra could last for 10 weeks at biologically relevant levels following delivery with recombinant AAV in the forelimb joints of horses (Watson et al., 2013). In a phase I clinical trial (NCT00617032), 15 subjects (aged ≥18 years) with inflammatory arthritis (14 with RA and 1 with ankylosing spondylitis) received a single IA injection of rAAV-2 containing a TNF immunoglobulin Fc fusion gene (rAAV2-TNFR:Fc). The result showed that rAAV2-TNFR:Fc appears to be safe and well tolerated in patients not systemically taking TNF-α antagonists (Mease et al., 2009). Furthermore, a phase I/II clinical trial (NCT00126724) on 127 patients (aged 18–75 years) demonstrated that greater improvement in patients treated with rAAV2-TNFR:Fc compared to placebo patients (Mease et al., 2010). Meanwhile, another phase I study (NCT02790723) using AAV is still in progress and will evaluate the effects of IL-1Ra expression on the OA phenotype (Bellavia et al., 2018).

Among the numerous target genes, the expressions of GFs and anti-inflammatory cytokines are of interest to researchers and are being assessed in clinical trials (see Table 3). As described aforementioned, IL-1 and TNF-α receptor antagonists are the candidate transgenes with an anti-inflammatory action that have been applied in clinical trials (Mease et al., 2009; Bellavia et al., 2018). TGF-β1 is a growth factor that can augment the ability of chondrogenesis of MSCs. Several preliminary studies have confirmed the effects of TGF-β1 in the repair of cartilage defects and impeding the chondrocytes hypertrophy (van Caam et al., 2015; Xie et al., 2016). TGF-β1 is also the exclusive gene currently being tested in clinical trials for OA treatment (Ondresik et al., 2017). Noh et al. conducted a pre-clinical study to evaluate the efficacy, biodistribution, and safety of single IA injection of the cell mixture (3:1 ratio of genetically unmodified and TGF-β1-secreting human chondrocytes, TG-C) in SCID mice, rabbits, and goats with damaged joints. The results showed that the mixture was tolerated well in all of the species, and cartilage regeneration was present in defects of rabbits and goats (Noh et al., 2010). After these positive pre-clinical results, a phase I clinical trial (NCT00599248) performed on 12 subjects with advanced knee OA suggested that TG-C contributed to an improvement of OA symptoms as well as minor injection site reactions (Ha et al., 2012). Concomitantly, a phase II clinical study carried out on 102 patients (NCT01221441) with knee OA, indicated that TG-C treated patients had positive effects on function elevation and pain mitigation compared to placebo at 1-year follow-up (Cherian et al., 2015). Notably, an ex vivo TGF-β1 gene therapy was authorized in Korea for IA injection of knee joints with moderate-to-severe OA. The product (Invossa™) has received marketing approval in Korea and a phase III clinical trial is expected to begin shortly in the United States (Evans et al., 2018). Other genes, like IGF-1, BMPs, cell signaling protein iHH, ECM component (COMP), and integrin β1, are still in their infancy (Bellavia et al., 2018; Tendulkar et al., 2019). Gene combinations are also of interest, and co-infection of IL-1Ra and IGF-1 has a positive effect on repairing cartilage defects in vivo (Morisset et al., 2007).

Gene therapy offers a novel approach to address the issue of transferring exogenous pharmaceuticals into joints topically and durably. Nevertheless, safety and effectiveness are still hurdles for clinical application. Some authors expressed their concerns about the safety of viral vectors, in particular, after the occurrence of severe adverse events such as leukemia and death (Frank et al., 2009). Although these events were not correlated with viral vectors, the proper monitoring for further clinical application needed to be highlighted.