CCL2 Promotes Novel Coronavirus-Mediated Inflammatory Responses in Macrophages.

PURPOSE: The hyperinflammatory response is one of the main complications associated with novel coronavirus disease 2019 (COVID-19), and there is no effective treatment for cytokine storm. Therefore, it is important to investigate the key genes associated with severity of the disease. METHODS: In this study, we used a microarray data set to analyze the key genes associated with severe illness in patients with COVID-19. The proportion of immune cells was determined using the CIBERSORT algorithm. The key genes were further verified by detecting the levels of cytokines and chemokines in the serum of patients. Additionally, macrophages were stimulated with SARS-CoV-2 spike protein and chemokine ligand (CCL) 2. The expression of cytokines, ERK1/2, and NF-κB in macrophages was detected. RESULTS: Four hub genes were identified. Among them, C-C motif chemokine receptor 2 (CCR2) was an upregulated hub gene, while killer cell lectin-like receptor subfamily K member 1 (KLRK1), macrophage colony-stimulating factor receptor (CSF1R), and CD3D human recombinant protein (CD3D) were downregulated genes. Immune cell type identification found that the proportion of monocytes was higher in patients with severe COVID-19 than that in controls. Moreover, levels of CCL2 were significantly higher in patients with COVID-19. When stimulated with SARS-CoV-2 S protein and CCL2, macrophages secreted more inflammatory cytokines. The expression level of ERK1/2 was elevated. CONCLUSIONS: These results suggested that S protein and CCL2 may mediate macrophage inflammatory responses through the ERK1/2 signaling pathway. This study provides a basis for clinical treatment and improves the prognosis of critically ill patients with COVID-19.

Decrease of 5-hydroxymethylcytosine in hepatitis B virus-related hepatocellular carcinoma: A cross-sectional study.

Many epigenetic studies had found the decrease of 5-hydroxymethylcytosine (5-hmC) in various tumor tissues. However, limited information is available for hepatitis B virus-related hepatocellular carcinoma (HBV-related HCC). The present study aimd to investigate whether the decrease also existed in tumor tissues of HBV-related HCC and, if possible, to disclose its mechanism. We used immunohistochemistry and Image Pro Plus 6.0 Image Analysis Software to quantify the expression of 5-hmC, 5-methylcytosine, 10-eleven translocation (TET), isocitrate dehydrogenase (IDH) in pathological sections of tumor tissues and its para cancerous tissues of 40 HBV-related HCC patients. Our results showed that 5-hmC was decreased while 5-methylcytosine was increased in tumor tissues. We also detected TET1 and IDH2 were decreased in the tumor tissues and the decrease were positively correlated with the 5-hmC. The results suggested that the deficiency of 5-hmC was an epigenetic characteristic of HBV-related HCC and was mainly caused by the decrease of TET1 and IDH2.

Diagnostic value and characteristic analysis of serum nucleocapsid antigen in COVID-19 patients.

Background: To date, several types of laboratory tests for coronavirus disease 2019 (COVID-19) diagnosis have been developed. However, the clinical importance of serum severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid antigen (N-Ag) remains to be fully elucidated. In this study, we sought to investigate the value of serum SARS-CoV-2 N-Ag for COVID-19 diagnosis and to analyze N-Ag characteristics in COVID-19 individuals. Methods: Serum samples collected from 215 COVID-19 patients and 65 non-COVID-19 individuals were used to quantitatively detect N-Ag via chemiluminescent immunoassay according to the manufacturer's instructions. Results: The sensitivity and specificity of the N-Ag assay were 64.75% (95% confidence interval (95% CI) [55.94-72.66%]) and 100% (95% CI [93.05-100.00%]), respectively, according to the cut-off value recommended by the manufacturer. The receiver operating characteristic (ROC) curve showed a sensitivity of 100.00% (95% CI [94.42-100.00%]) and a specificity of 71.31% (95% CI [62.73-78.59%]). The positive rates and levels of serum SARS-CoV-2 N-Ag were not related to sex, comorbidity status or disease severity of COVID-19 (all P < 0.001). Compared with RT‒PCR, there was a lower positive rate of serum N-Ag for acute COVID-19 patients (P < 0.001). The positive rate and levels of serum SARS-CoV-2 N-Ag in acute patients were significantly higher than those in convalescent patients (all P < 0.001). In addition, the positive rate of serum SARS-CoV-2 N-Ag in acute COVID-19 patients was higher than that of serum antibodies (IgM, IgG, IgA and neutralizing antibodies (Nab)) against SARS-CoV-2 (all P < 0.001). However, the positive rate of serum SARS-CoV-2 N-Ag in convalescent COVID-19 patients was significantly lower than that of antibodies (all P < 0.001). Conclusion: Serum N-Ag can be used as a biomarker for early COVID-19 diagnosis based on appropriate cut-off values. In addition, our study also demonstrated the relationship between serum N-Ag and clinical characteristics.

Exploring the relationships between hormone receptor, HER2 status, and bone involvement in the first distant metastases of in Chinese breast cancer patients who lacked HER2 targeted therapy.

BACKGROUND: This retrospective study explored the relationship between hormone receptor (HR), human epidermal growth factor receptor 2 (HER2) status, and bone involvement in the first distant metastases (DM) of Chinese breast cancer (BC) patients who lacked the HER2 targeted therapy. Such therapy was rarely received due to its lag approval or high cost in China compared with the developed countries. METHODS: All eligible women with primary unilateral stage I - III BC and first DM diagnosed in 2008-2018 at one cancer center were identified for enrollment. Based on chart records, a full or no/partial compliance status of endocrine therapy (ET) was assigned for HR-positive patients. Multivariate logistic regression was used to estimate the adjusted odd ratio (aOR), its 95%CI and p value. RESULTS: Four hundred eighteen patients had an average age of 50.7 years and median disease-free survival of 27.1 months at DM. Bone, lung, liver and brain metastasis rates in patients were 55.7%, 34.7%, 33.0% and 8.1%, respectively. Compared to HR-negative patients, HR-positive patients with the full and non/partial compliance of ET were significantly associated with higher risk of bone involvement with an aOR of 2.329 (1.316 - 1.741, p = 0.004) and 2.317 (1.330 - 4.036, p = 0.003), respectively. No difference of such risk was found between the two groups of ET compliance (p = 0.984) nor between HER2-negative and HER2-positive patients (aOR 0.827, p = 0.431). Stratified analyses further indicated that HR-positive was associated with bone involvement only in HER2-negative BC patients (p = 0.006-0.015). CONCLUSIONS: HR-positive tumors are significantly associated with bone involvement in HER2-negative metastatic BC patients. ET does not appear to impact this association. HER2 status per se is not associated with such risk.

A Novel Rat Model of Patellofemoral Osteoarthritis Due to Patella Baja, or Low-Lying Patella.

BACKGROUND Patella baja, or patella infera, consists of a low-lying patella that results in a limited range of motion, joint pain, and crepitations. Patellofemoral joint osteoarthritis (PFJOA) is a subtype OA of the knee. This study aimed to develop a reproducible and reliable rat model of PFJOA. MATERIAL AND METHODS Three-month-old female Sprague-Dawley rats (n=24) included a baseline group (n=8) that were euthanized at the beginning of the study. The sham group (n=8), and the patella ligament shortening (PLS) group (n=8) were euthanized and evaluated at ten weeks. The PLS model group (n=8) underwent insertion of a Kirschner wire under the patella tendon to induce patella baja. At ten weeks, the sham group and the PLS group were compared using X-ray imaging, macroscopic appearance, histology, immunohistochemistry, TUNEL staining for apoptosis, and micro-computed tomography (micro-CT). The patella height was determined using the modified Insall-Salvati (MIS) ratio. RESULTS The establishment of the rat model of patella baja in the PLS group at ten weeks was confirmed by X-ray. In the PLS group, patella volume, sagittal length, and cross-sectional area were significantly increased compared with the sham group. The PFJ showed typical lesions of OA, confirmed macroscopically and histologically. Compared with the sham group, in the rat model of PFJOA, there was increased cell apoptosis, and immunohistochemistry showed increased expression of biomarkers of osteoarthritis, compared with the sham group. CONCLUSIONS A rat model of PFJOA was developed that was confirmed by changes in cartilage and subchondral bone.

Salmon calcitonin exerts better preventive effects than celecoxib on lumbar facet joint degeneration and long-term tactile allodynia in rats.

OBJECTIVE: To evaluate and compare the effects of salmon calcitonin (sCT) and celecoxib (CLX) on cartilage, subchondral bone and tactile allodynia in a rat model of lumbar facet joint (FJ) osteoarthritis (OA). METHOD: Forty 3-month-old male Sprague-Dawley rats were randomly divided into four groups: 30 received surgical collagenase (type II) injections in the right L3-L6 facet joints followed by 8 weeks of treatment with normal saline, CLX or sCT, and the other 10 received sham surgery. Tactile allodynia, changes of cartilage and subchondral bone of the L4-L5 FJs, and serum biomarkers were analyzed for all rats. RESULTS: Both sCT and CLX ameliorated cartilage lesions, significantly increased aggrecan expression and decreased caspase-3 expression. sCT also decreased the expression of a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS-4). According to the micro-computed tomography (micro-CT) analysis, sCT significantly improved microarchitecture parameters of subchondral bone and micro-CT score; and inhibited articular process hypertrophy. CLX showed better antihyperalgesic effects than sCT on days 3 and 7 postoperatively despite no statistical differences, whereas sCT possessed better analgesic effects than CLX on days 42 and 56. Besides, the sCT treatment reduced the elevated cartilage oligomeric matrix protein (COMP) concentration in rats injected with collagenase (type II). CONCLUSIONS: Both sCT and CLX exerted preventive effects on FJ OA caused by collagenase (type II), but sCT showed more protective effects, particularly on maintaining cartilage metabolism, restraining the deterioration of the subchondral bone microarchitecture and tactile allodynia, and reducing serum COMP concentrations.

Orally administered simvastatin partially preserves lumbar vertebral bone mass but not integrity of intervertebral discs in ovariectomized rats.

The present study aimed to investigate the effect of orally administered simvastatin on lumbar vertebral bone mass and intervertebral disc (IVD) degeneration in ovariectomized (OVX) rats. A total of 30 female Sprague-Dawley (SD) rats were subjected to either bilateral ovariectomy (n=20) or sham surgery (n=10). After 12 weeks, the OVX rats were orally administered either saline vehicle (OVX + V group; n=10), or 5 mg/kg/day simvastatin (OVX + SIM group; n=10). Following 12 weeks of treatment, necropsy was conducted and bone mineral density (BMD) was determined in the L5-6 vertebrae. Furthermore, the microstructure and biomechanical properties of the L3 vertebrae were detected by micro-computed tomography and compression testing, respectively. The L5-6 vertebrae were analyzed by measurement of IVD height, observation of histological changes by van Gieson staining, and evaluation of collagen-II (col-II), aggrecan (AGG) and collagen I (col-I) expression by immunohistochemical analysis. Rats in the OVX+V group had lower BMD, bone volume/trabecular volume ratio, maximum load and elastic modulus than the sham group. Rats in the OVX + SIM group had higher BMD and biomechanical strength values than the rats in the OVX+V group. Histological analysis showed that the OVX + V and OVX + SIM groups exhibited significantly higher disc degeneration scores and significantly lower IVD height than the sham group. Immunohistochemical analysis revealed lower expression levels of col-II and AGG, but higher levels of col-I in the annulus fibrosis and endplate in OVX+V rats compared with the sham group. The OVX + SIM group exhibited levels of col-II, AGG and col-I expression comparable with those of OVX+V rats, with the exception of an upregulation of col-II expression in the annulus fibrosis. These data demonstrate that simvastatin treatment partially prevented bone loss and the deterioration of biomechanical properties of lumbar vertebrae, but not the progression of IVD degeneration in OVX rats.

Effects of alendronate on lumbar intervertebral disc degeneration with bone loss in ovariectomized rats.

BACKGROUND CONTEXT: Osteoporosis adversely affects disc degeneration cascades, and prophylactic alendronate (ALN) helps delay intervertebral disc degeneration (IDD) in ovariectomized (OVX) rats. However, there remains no information regarding whether ALN affects IDD with bone loss. PURPOSE: This study aimed to observe the effects of ALN on degenerative discs with bone loss induced by OVX in rats. STUDY DESIGN: This study used controlled in vivo experiments in rodents. METHODS: Thirty female Sprague-Dawley rats were randomly assigned to undergo sham surgery (n=10) or OVX surgery (n=20); 3 months later, the OVX animals were injected with either ALN (OVX+ALN, 15 µg/kg/2w, n=10) or normal saline (OVX+vehicle treatment [V], n=10). At 3 months after the ALN intervention, van Gieson staining and immunohistochemistry were used to investigate histologic and metabolic changes in the discs. Bone mineral density (BMD), micro-computed tomography, and biomechanical tests were conducted to determine the biological properties of the vertebrae. RESULTS: The OVX+ALN group exhibited significantly reduced morphologic degenerative alterations in both the nucleus pulposus and annulus fibrosus, with a markedly lower IDD score than that of the OVX+V group. The OVX+ALN samples showed increased disc height and decreased cartilage end plate thickness and bony area compared with the OVX+V group. Compared with saline, ALN administration markedly inhibited the type I collagen, matrix metalloprotease (MMP)-1, and MMP-13 expression levels while increasing the type II collagen and aggrecan expression levels in the disc matrix. Compared with the OVX+V group, OVX+ALN vertebrae revealed significantly enhanced BMD with increased biomechanical strength, as well as increased percent bone volume and trabecular thickness. CONCLUSIONS: ALN has favorable effects on disc degeneration with bone loss and helps to alleviate IDD while enhancing the biological and mechanical properties of vertebrae and end plates.

Strontium ranelate causes osteophytes overgrowth in a model of early phase osteoarthritis.

BACKGROUND: Osteoarthritis (OA) involves cartilage changes as well as modifications of subchondral bone and synovial tissues. Strontium ranelate (SR), an anti-osteoporosis compound, which is currently in phase III clinical trial for treatment of OA. Evidences suggest that SR preferably deposited in osteophyte, other than in subchondral bone in early phase of OA. This phenomenon raises concern about its utility for OA treatment as a disease-modifying drug. To evaluate the effect of SR on cartilage, subchondral bone mass and subchondral trabecular bone structure in medial meniscectomized (MNX) guinea pigs. METHOD: Thirty-six 3-month-old male Dunkin Hartley albino guinea pigs received either sham or medial meniscectomy operations. One week after the procedure, meniscectomized animals began 12 weeks of SR (625 mg/kg, daily) treatment by oral gavage for MNX + SR group, or normal saline for MNX + V group. All animals were euthanized 12 weeks later, cartilage degeneration and subchondral bone micro-architecture was analyzed. RESULTS: Both OARSI scores (P = 0.523 for marcoscopic scores, P = 0.297 for histological scores) and Cartilage thickness (P = 0.335) in MNX + SR group were comparable to MNX + V group. However, osteophyte sizes were larger in MNX + SR group (P = 0.014), and collapsed osteophytes in MNX + SR group (7 by 12) were significantly more than in MNX + V group (1 by 12) (P = 0.027), while immunohistochemistry indicates catabolic changes in osteophyte/plateau junction. Micro-CT analysis showed bone mineral density (BMD) (P = 0.001), bone volume fraction (BV/TV) (P = 0.008), trabecular spacing (Tb.Sp) (P = 0.020), trabecular thickness (Tb.Th) (P = 0.012) and structure model index (SMI) (P = 0.005) levels to be significantly higher in the MNX + SR group than in the MNX + V group. CONCLUSIONS: SR (625 mg/kg/day) did not protect cartilage from degeneration in MNX guinea pigs but subchondral bone was significantly enhanced. In early phase OA, SR administration causes osteophyte overgrowth, which may be related to incorporation into mineralizing osteophytes. This adverse effect is important for future studies of SR in OA.

Are estrogen-related drugs new alternatives for the management of osteoarthritis?

Osteoarthritis (OA) is a chronic degenerative disease involving multiple physiopathological mechanisms. The increased prevalence of OA after menopause and the presence of estrogen receptors in joint tissues suggest that estrogen could help prevent development of OA. This review summarizes OA research with a focus on the effects of estrogen and selective estrogen receptor modulators (SERMs). Preclinical studies and clinical trials of estrogen therapy have reported inconsistent results. However, almost all studies assessing SERM treatment have obtained more consistent and favorable effects in OA with a relatively safety and tolerability profiles. At present, some SERMs including raloxifene and bazedoxifene have been approved for the treatment of osteoporosis. In summary, estrogen-related agents may exert both a direct effect on subchondral bone and direct and/or indirect effects upon the surrounding tissues, including the articular cartilage, synovium, and muscle, to name a few. Estrogen and SERMs may be particularly favorable for postmenopausal patients with early-stage OA or osteoporotic OA, a phenotype defined by reduced bone mineral density related to high remodeling in subchondral bone. At present, no single drug exists that can prevent OA progression. Although estrogen-related drugs provide insight into the continued work in the field of OA drug administration, further research is required before SERMs can become therapeutic alternatives for OA treatment.

[Research progress of effects of strontium ranelate on osteoarthritis].

OBJECTIVE: To review the research progress focused on the effects of strontium ranelate (SR) on osteoarthritis. METHODS: The relevant literature about the effects and mechanism of SR intervening osteoarthritis in recent years was extensively reviewed and comprehensively analyzed. RESULTS: SR not only could improve the microenvironment of bone metabolism in articular cartilage with osteoarthritis, promote activity of osteoblasts, and inhibit activity of osteoclasts, but also could adjust the expression of key proteases which affect cartilage formation, and therefore it has a potential protective effect on subchondral bone during the progression of osteoarthritis cartilage. CONCLUSION: SR is expected to become a drug of osteoarthritis disease remission, but further studies are needed to clarify the mechanism of SR in osteoarthritis, and finally confirm the best application dosage of SR in osteoarthritis treatment.