Jianmin Li

Increased Frequencies of Th22 Cells as well as Th17 Cells in the Peripheral Blood of Patients with Ankylosing Spondylitis and Rheumatoid Arthritis

Background T-helper (Th) 22 is involved in the pathogenesis of inflammatory diseases. The roles of Th22 cells in the pathophysiological of ankylosing spondylitis (AS) and rheumatoid arthritis (RA) remain unsettled. So we examined the frequencies of Th22 cells, Th17 cells and Th1 cells in peripheral blood (PB) from patients with AS and patients with RA compared with both healthy controls as well as patients with osteoarthritis. Design and Methods We studied 32 AS patients, 20 RA patients, 10 OA patients and 20 healthy controls. The expression of IL-22, IL-17 and IFN-γ were examined in AS, RA, OA patients and healthy controls by flow cytometry. Plasma IL-22 and IL-17 levels were examined by enzyme-linked immunosorbent assay. Results Th22 cells, Th17 cells and interleukin-22 were significantly elevated in AS and RA patients compared with OA patients and healthy controls. Moreover, Th22 cells showed positive correlation with Th17 cells as well as interleukin-22 in AS and RA patients. However, positive correlation between IL-22 and Th17 cells was only found in AS patients not in RA patients. In addition, the percentages of both Th22 cells and Th17 cells correlated positively with disease activity only in RA patients not in AS patients. Conclusions The frequencies of both Th22 cells and Th17 cells were elevated in PB from patients with AS and patients with RA. These findings suggest that Th22 cells and Th17 cells may be implicated in the pathogenesis of AS and RA, and Th22 cells and Th17 cells may be reasonable cellular targets for therapeutic intervention.

Diallyl trisulfide inhibits proliferation, invasion and angiogenesis of osteosarcoma cells by switching on suppressor microRNAs and inactivating of Notch-1 signaling

Notch signaling pathway plays critical roles in human cancers, including osteosarcoma, suggesting that the discovery of specific agents targeting Notch would be extremely valuable for osteosarcoma. Our previous studies have shown that diallyl trisulfide (DATS) inhibits proliferation of osteosarcoma cells by triggering cell cycle arrest and apoptosis in vitro. However, the underlying mechanism is still unclear. In this study, we found that DATS suppressed cell survival, wound-healing capacity, invasion and angiogenesis in osteosarcoma cells. These effects were associated with decreased expression of Notch-1 and its downstream genes, such as vascular endothelial growth factor and matrix metalloproteinases, as well as increased expression of a panel of tumor-suppressive microRNAs (miRNAs), including miR-34a, miR-143, miR-145 and miR-200b/c that are typically lost in osteosarcoma. We also found that reexpression of miR-34a and miR-200b by transfection led to reduced expression of Notch-1, resulting in the inhibition of osteosarcoma cell proliferation, invasion and angiogenesis. These results clearly suggest that DATS inhibited osteosarcoma growth and aggressiveness via a novel mechanism targeting a Notch-miRNA regulatory circuit. Our data provide the first evidence that the downregulation of Notch-1 and reexpression of miRNAs by DATS may be an effective approach for the treatment of osteosarcoma.

Curcumin inhibits proliferation and invasion of osteosarcoma cells through inactivation of Notch‐1 signaling

The Notch signaling pathway plays critical roles in human cancers, including osteosarcoma, suggesting that the discovery of specific agents targeting Notch would be extremely valuable for osteosarcoma. Curcumin, a naturally occurring phenolic compound found in curcuma longa, has been shown to inhibit proliferation and induce apoptosis of osteosarcoma cells inu2003vitro and tumor growth in xenotransplant or orthotransplant models. However, the precise molecular mechanisms by which curcumin exerts its antitumor activity remain unclear. Here we used multiple molecular approaches, such as the 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide assay, the invasion assay, gene transfection, real‐time RT‐PCR, western blot and gelatin zymography, to investigate whether the downregulation of Notch‐1 contributes to curcumin‐induced inhibition of proliferation and invasion in osteosarcoma cells. The results showed that curcumin caused marked inhibition of osteosarcoma cell growth and G2/M phase cell cycle arrest. This was associated with concomitant attenuation of Notch‐1 and downregulation of its downstream genes, such as matrix metalloproteinases, resulting in the inhibition of osteosarcoma cell invasion through Matrigel. We also found that specific downregulation of Notch‐1 via small‐interfering RNA prior to curcumin treatment resulted in enhanced inhibition of cell growth and invasion. These results suggest that antitumor activity of curcumin is mediated through a novel mechanism involving inactivation of the Notch‐1 signaling pathway. Our data provide the first evidence that the downregulation of Notch‐1 by curcumin may be an effective approach for the treatment of osteosarcoma.

Limb sparing surgery for bone tumours of the shoulder girdle: the oncological and functional results

It is a great challenge to spare the upper limb with a malignant or invasive benign bone tumour of the shoulder girdle. We retrospectively analysed 35 patients with bone tumours of the shoulder girdle treated with various limb salvage procedures. The tumours included 25 primary malignancies, three metastases and seven giant cell tumours which involved the proximal humerus in 21 patients, scapula in 12 and clavicle in two. The reconstruction procedures included eight prosthetic replacements, four devitalised tumorous bone grafts, three osteoarticular allografts, two autogenous fibular grafts, one intramedullary cemented nail, three Tikhoff-Linberg procedures, two replantation of shortened arms, and four humeral head suspensions. Six partial scapulectomies and two lateral clavicectomies needed no bone reconstruction. With an average follow-up of 71xa0months, local recurrences occurred in four cases and systemic metastases in six. Nine patients died and 23 remained disease free. The five year Kaplan-Meier survival rate of 28 patients with malignancies was 69.5%. The average Musculoskeletal Tumour Society (MSTS) functional score was 77% (range 40–100%) in all patients.

Long non-coding RNA tumor suppressor candidate 7 functions as a tumor suppressor and inhibits proliferation in osteosarcoma.

Osteosarcoma is the most common malignant tumor of bone. Recent studies have proven long non-coding RNAs (lncRNAs) play important roles in the tumorigenesis and progression of cancer. However, few lncRNAs have been investigated in osteosarcoma. Here, we reported a novel lncRNA, tumor suppressor candidate 7 (TUSC7), was significantly downregulated in osteosarcoma tissues compared with paired non-tumor tissues and low expression of TUSC7 indicated poor survival (HRu2009=u20090.313, 95xa0% confidence interval (CI) 0.092–0.867) of osteosarcoma patients. Further analysis revealed that loss copy number of TUSC7 was correlated with low expression of TUSC7, and additionally, loss of TUSC7 copy number also indicated poor prognosis (HRu2009=u20093.994, 95xa0% CI 1.147–13.91) of osteosarcoma patients. Two osteosarcoma cell lines, HOS and MG63, were utilized to investigate biological function of TUSC7. Cell counting kit 8 (CCK-8) assay revealed that after silence of TUSC7, cell proliferation ability increased and the colony formation ability also increased. Further results showed that cell cycle was not affected by treatment of si-TUSC7, while the percentage of apoptotic cells decreased. Western blot showed that after silence of TUSC7, the proapoptotic Bcl2 expression was downregulated. Finally, we established xenograft tumor models in nude mice with MG63 cells. Compared with negative control group, silence of TUSC7 significantly promoted tumor growth in vivo. Thus, we demonstrated that TUSC7 could be a potential tumor suppressor in osteosarcoma.

CD271+ Osteosarcoma Cells Display Stem-Like Properties

Cancer stem cell (CSC) theory has been proposed and verified in many cancers. The existence of osteosarcoma CSCs has been confirmed for many years and multiple surface markers have been employed to identify them. In this study, we identified CD271+ subpopulation of osteosarcoma displaying stem-like properties. CD271, known as the neural crest nerve growth factor receptor, is the marker of bone marrow mesenchymal stem cells (MSCs) and human melanoma-initiating cells. We discovered that CD271 was expressed differentially in diverse types of human osteosarcoma and stabilized cell lines. CD271+ osteosarcoma cells displayed most of the properties of CSC, such as self-renewal, differentiation, drug resistance and tumorigenicity in vivo. Nanog, Oct3/4, STAT3, DNA-PKcs, Bcl-2 and ABCG2 were more expressed in CD271+ cells compared with CD271− cells. Our study supported the osteosarcoma CSC hypothesis and, to a certain extent, revealed one of the possible mechanisms involved in maintaining CSCs properties.

Incorporation of Methotrexate in Calcium Phosphate Cement: Behavior and Release in Vitro and in Vivo

The application of antibiotic-loaded calcium phosphate bone cement in the treatment and prevention of osteomyelitis suggests that calcium phosphate cement could also be used as an anticancer drug carrier to reduce the local recurrence of bone tumors and systemic toxicities of chemotherapy. We added 0 to 400 mg of methotrexate to 40 g cement, with final methotrexate concentrations of 0% to 1% (weight/weight). The setting times, mechanical properties, microstructures, and in vitro methotrexate release kinetics of these methotrexate-calcium phosphate bone cement specimens were evaluated, along with in vivo methotrexate release kinetics in 24 rabbits. Methotrexate did not significantly alter the cement setting time. The compressive and tensile strengths of the methotrexate-calcium phosphate bone cement specimens were significantly less when 400 mg methotrexate was used, compared to control samples without methotrexate addition. Nevertheless, the cement remained compliant with the minimum requirements for clinical application. The scanning electron microscopy micrographs showed that the basic crystal structure did not alter. The methotrexate release kinetics in vitro and in vivo confirmed that methotrexate-calcium phosphate bone cement was a monolithic matrix system, with a burst effect in the initial stage and a sudden drop thereafter. Drug delivery in vivo was faster than in vitro. We estimated that the incorporated methotrexate could be continuously released over 2 to 4 months at a higher than minimum concentration. The methotrexate had no apparent toxicity on the host rabbits, even at the highest dose of methotrexate tested. The methotrexate-calcium phosphate bone cement system may be a potentially effective therapy for bone tumors in humans.

Diallyl trisulfide reverses drug resistance and lowers the ratio of CD133+ cells in conjunction with methotrexate in a human osteosarcoma drug-resistant cell subline

P-glycoprotein (P-gp) overexpression and tumor stem cells are thought to be important factors in the cross-resistance of cancer cells. There have been no studies on whether diallyl trisulfide (DATS) can reverse drug resistance in human osteosarcoma cells. In the present study, we demonstrated that DATS down-regulated P-gp expression and reversed drug resistance. DATS and methotrexate (MTX) decreased the ratio of drug-resistant human osteosarcoma cells positive for CD133 (a tumor stem cell marker). To the best of our knowledge, this is the first evidence that DATS can reverse drug resistance and lower the ratio of CD133+ cells in human osteosarcoma cells in conjunction with MTX.

Diallyl trisulfide induces osteosarcoma cell apoptosis through reactive oxygen species-mediated downregulation of the PI3K/Akt pathway.

Diallyl trisulfide (DATS) is a natural organosulfur compound isolated from garlic, and has been reported to possess anticancer activities. However, the cancer growth inhibitory effects and molecular mechanisms in human osteosarcoma cells have not been well studied. The present study demonstrated that DATS significantly reduced cell viability in a dose-xa0and time-dependent manner in MG63 and MNNG/HOS cells. DATS-induced G0/G1xa0phase arrest was found to correlate with a decrease in cyclinxa0D1 in concomitance with an increase in p21 and p27. DATS induced a marked increase in reactive oxygen species (ROS) levels and collapse of mitochondrial membrane potential (Δψm) in the osteosarcoma cells. DATS induced apoptosis in the MG63 and MNNG/HOS cells via inhibition of the PI3K/Akt signaling pathway and through the mitochondrial apoptotic pathway. The efficiency of DATS basically approached the efficacy of LY294002, a specific PI3K inhibitor. However, N-acetylcysteine (NAC), a general ROS scavenger, completely blocked the DATS-induced ROS increase, inhibition of the PI3K/Akt pathway and cell apoptosis. Overall, DATS has the potential to be developed as a new anticancer drug. The mechanisms of action involve the ROS-mediated downregulation of the PI3K/Akt pathway.

Targeting DNA-PKcs increased anticancer drug sensitivity by suppressing DNA damage repair in osteosarcoma cell line MG63

Many chemotherapy drugs exert anticancer effects through causing DNA damage, such as DNA topoisomerase inhibitor and platinum-containing drugs. DNA damage repair is an important mechanism of drug resistance which is responsible for metastasis and recurrence after chemotherapy. DNA-dependent protein kinase (DNA-PK) plays an important role in non-homology end joining (NHEJ) pathway. In this study, we aimed to determine whether DNA-PK catalytic subunit (DNA-PKcs) is expressed in osteosarcoma MG63 cell line and involved in drug resistance induced by DNA repair. We found that DNA-PKcs was expressed in osteosarcoma cell line MG63. The pDNA-PKcsT2609 was more expressed in cells treated with cisplatin (DDP) and etoposide (VP16). Down-regulation of DNA-PKcs produced higher sensitivity of MG63 cells to DDP or VP16 through increasing apoptosis and causing cell cycle arrest in the G1 phase. Our study supported that DNA-PKcs was involved in drug-induced DNA damage repair and related to chemosensitivity of osteosarcoma MG63 cells.