Monocytes and γδ T cells control the acute‐phase response to intravenous zoledronate: Insights from a phase IV safety trial

Aminobisphosphonates (NBPs) are used widely against excessive bone resorption in osteoporosis and Paget's disease as well as in metastatic bone disease and multiple myeloma. Intravenous NBP administration often causes mild to severe acute‐phase responses (APRs) that may require intervention with analgesics and antipyretics and lead to treatment noncompliance and nonadherence. We here undertook a phase IV safety trial in patients with osteoporosis to investigate the APR of otherwise healthy individuals to first‐time intravenous treatment with the NBP zoledronate. This study provides unique insight into sterile acute inflammatory responses in vivo, in the absence of confounding factors such as infection or cancer. Our data show that both peripheral γδ T cells and monocytes become rapidly activated after treatment with zoledronate, which ultimately determines the clinical severity of the APR. Our study highlights a key role for IFN‐γ in the zoledronate‐induced APR and identifies pretreatment levels of monocytes and central/memory Vγ9/Vδ2 T cells as well as their responsiveness to zoledronate in vitro as predictive risk factors for the occurrence of subclinical and clinical symptoms. These findings have diagnostic and prognostic implications for patients with and without malignancy and are relevant for Vγ9/Vδ2 T‐cell–based immunotherapy approaches. © 2012 American Society for Bone and Mineral Research. © 2013 American Society for Bone and Mineral Research.     

Reconstitution of T and NK cells after haploidentical hematopoietic cell transplantation using αβ T cell–depleted grafts and the clinical implication of γδ T cells

To investigate reconstitution of T and NK cells after αβ T lymphocyte–depleted haploidentical hematopoietic cell transplantation (HHCT) and the clinical implications of γδ T cells, we analyzed 50 pediatric patients who received 55 HHCTs using αβ T cell–depleted grafts. The number of CD3+ T cells and CD8+ T cells recovered rapidly and reached donor levels at days 180 and 60, respectively. Recovery of NK cells was rapid, and the median of NK cells at day 14 was comparable to the donor level. At day 14, median percentage of γδ T lymphocytes was 70.5%. After day 14, the percentage of γδ T cells gradually decreased, while the percentage of αβ T cells gradually increased. Patients with a low percentage (≤21%) of γδ T cells at day 30 had significantly higher incidence of cytomegalovirus (CMV) reactivation compared to patients with a high percentage (>70%) of γδ T cells (P < .01). In patients with acute leukemia, patients with high percentage of γδ T cells at day 30 showed significantly higher relapse‐free survival compared to those with low percentage of γδ T cells (P = .02). Data suggest that early recovery of γδ T cells decreases the risk of CMV reactivation and leukemia relapse.     

Characterization of γδ T cell subsets in organ transplantation

γδ T cells are innate‐type lymphocytes that preferentially act as regulators of local effector immune responses. Recent reports found an altered distribution of the two main subpopulations of blood γδ T cells (Vδ1 and Vδ2) in operationally tolerant liver transplant recipients. Based on this, γδ T cells subset quantification was proposed as a biomarker of immunologic risk in liver transplantation. The specific characteristics of γδ T cell subsets in transplantation remain however unknown. We have investigated here the phenotype, repertoire and functional properties of γδ T cell subsets in a large population of allograft recipients. Our results indicate that alterations in the γδ T cell compartment are not restricted to tolerant liver recipients. In fact, most immunosuppressed liver and kidney recipients also display an enlarged peripheral blood γδ T cell pool mainly resulting from an expansion of Vδ1 T cells exhibiting an oligoclonal repertoire and different phenotypic and cytokine production traits than Vδ2 T cells. We propose that persistent viral infections are likely to contribute to these alterations. Our data provide novel insight in the biology of γδ T cells and a rationale for exploring these lymphocytes in more depth into the pathogenesis of viral infections in transplantation.     

An O‐Glycosylation of Fibronectin Mediates Hepatic Osteodystrophy Through α4β1 Integrin

Patients with cholestatic liver disease experience increased fracture risk. Higher circulating levels of a fibronectin isoform called oncofetal fibronectin (oFN) were detected in a subset of such patients. Administering this isoform to mice suppresses osteoblast differentiation and diminishes bone mineral density in vivo, suggesting it is responsible for bone loss in cholestatic liver disease. The aim of this study was to define the mechanism by which oFN affects osteoblast function and evaluate possible modifiers in experimental hepatic osteodystrophy. The fibronectin isoform oFN is characterized by the presence of various glycosylations. In line with this, adding oFN that underwent enzymatic O‐deglycosylation to osteoblasts normalized nodule formation in vitro. Of three possible O‐glycosylation sites in oFN, only a mutation at AA 33 of the variable region or binding of this glycosylated site with an antibody normalized osteoblast differentiation. Because the responsible site is located in the variable region of fibronectin, which binds to α4β1 or α4β7 integrins, these integrins were evaluated. We show that integrin α4β1 mediates the inhibitory effect of oFN both in vitro as well as in vivo. In a hepatic osteodystrophy mouse model, we demonstrate that liver fibrosis is associated with increased circulating oFN and diminished BMD. In addition, trabecular bone loss induced by oFN injection or fibrosis induction could be prevented by either administering an antibody that binds to α4 integrin (PS/2) or the CS1 peptide, which contains a binding site for α4β1 integrin. In summary, oFN inhibits osteoblast activity. This is because of an O‐glycosylation in the variable region that results in decreased integrin‐mediated signaling. This deleterious effect can be thwarted by binding α4β1 integrin. Thus, we have characterized the defect and the receptor mediating bone loss in patients with hepatic osteodystrophy and evaluated possible therapeutic interventions in a murine model. © 2016 American Society for Bone and Mineral Research.     

Human Osteoclasts Are Inducible Immunosuppressive Cells in Response to T cell–Derived IFN‐γ and CD40 Ligand In Vitro

Osteoclasts (OCs) are bone resorbing cells whose activity can be regulated by activated T cells and their cytokines. However, the immune function of OCs is largely unknown. In this study, we found that as bystanders, human OCs effectively suppressed T‐cell proliferation induced by allogeneic, microbial antigenic, and T‐cell receptor stimuli in vitro. Mechanism studies revealed that T cell–derived IFN‐γ and CD40 ligand (CD40L) induced the expression of indoleamine 2,3‐dioxygenase (IDO) in OCs, which mediated the immunosuppressive function on T‐cell proliferation through depleting tryptophan. Neutralizing IFN‐γ and blocking CD40L, or silencing or inhibiting IDO in OCs restored T‐cell proliferation in the presence of OCs. Our data reveal a novel function of human OCs as inducible immunosuppressive cells, and a feedback loop between OCs and activated T cells. Thus, this study provides new insight into the mechanism of the immunosuppressive function of OCs, and may be helpful for developing novel therapeutic strategies for human diseases involving both the bone and immune systems. © 2014 American Society for Bone and Mineral Research.     

Indirubin‐3′‐Oxime Reverses Bone Loss in Ovariectomized and Hindlimb‐Unloaded Mice Via Activation of the Wnt/β‐Catenin Signaling

Osteoporosis is a major global health issue in elderly people. Because Wnt/β‐catenin signaling plays a key role in bone homeostasis, we screened activators of this pathway through cell‐based screening, and investigated indirubin‐3′‐oxime (I3O), one of the positive compounds known to inhibit GSK3β, as a potential anti‐osteoporotic agent. Here, we show that I3O activated Wnt/β‐catenin signaling via inhibition of the interaction of GSK3β with β‐catenin, and induced osteoblast differentiation in vitro and increased calvarial bone thickness ex vivo. Intraperitoneal injection of I3O increased bone mass and improved microarchitecture in normal mice and reversed bone loss in an ovariectomized mouse model of age‐related osteoporosis. I3O also increased thickness and area of cortical bone, indicating improved bone strength. Enhanced bone mass and strength correlated with activated Wnt/β‐catenin signaling, as shown by histological analyses of both trabecular and cortical bones. I3O also restored mass and density of bone in hindlimb‐unloaded mice compared with control, suspended mice, demonstrating bone‐restoration effects of I3O in non‐aged–related osteoporosis as well. Overall, I3O, a pharmacologically active small molecule, could be a potential therapeutic agent for the treatment and prevention of osteoporosis. © 2014 American Society for Bone and Mineral Research.     

Thymosin β4 administration enhances fracture healing in mice

Thymosin β₄ (Tβ₄) is a regenerative peptide that we hypothesized would promote healing of fractured bone. Mice received a bilateral fibular osteotomy and were given i.p. injections of either Tβ₄ (6 mg/kg) or saline. Calluses from saline‐ and Tβ₄‐treated mice were analyzed for: (1) biomechanical properties and (2) composition using micro‐computed tomography (µCT) and histomorphometry. Biomechanical analysis showed that Tβ₄‐treated calluses had a 41% increase in peak force to failure (p < 0.01) and were approximately 25% stiffer (p < 0.05) than saline‐treated controls. µCT analysis at 21 days post‐fracture showed that the fractional volume of new mineralized tissue and new highly mineralized tissue were respectively 18% and 26% greater in calluses from Tβ₄‐treated mice compared to controls (p < 0.01; p < 0.05, respectively). Histomorphometry complemented the µCT data; at 21 days post‐fracture, Tβ₄‐treated calluses were almost 23% smaller (p < 0.05), had nearly 47% less old cortical bone (p < 0.05) and had a 31% increase in new trabecular bone area/total callus area fraction compared with controls (p < 0.05). Our finding of enhanced biomechanical properties of fractures in mice treated with Tβ₄ provides novel evidence of the therapeutic potential of this peptide for treating bone fractures. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:1277–1282, 2014.     

Pretransplant Interferon‐γ Secretion by CMV‐Specific CD8+ T Cells Informs the Risk of CMV Replication After Transplantation

In this prospective study we analyzed pretransplant interferon‐γ secretion by cytomegalovirus (CMV)‐specific CD8+ T cells to assess its possible utility in determining the risk of CMV replication after solid organ transplantation. A total of 113 lung and kidney transplant patients were enrolled in the study but only 55 were evaluable. All CMV‐seronegative recipients were pretransplant “nonreactive” (IFNγ <0.2 IU/mL) (11/11), whereas 30/44 (68.2%) CMV‐seropositive (R+) recipients were “reactive” (IFNγ ≥0.2 IU/mL) and 14/44 (31.8%) were “nonreactive”. In the R(+) “nonreactive” group, 7/14 (50%) developed posttransplant CMV replication, whereas the virus replicated only in 4/30 (13.3%) of the R(+) “reactive” patients (p = 0.021). According to the best multivariate model, pretransplant “nonreactive” recipients receiving an organ from a CMV‐seropositive donor had a 10‐fold increased risk of CMV replication compared to pretransplant “reactive” recipients (adjusted OR 10.49, 95% CI 1.88–58.46). This model displayed good discrimination ability (AUC 0.80) and calibration (Hosmer–Lemeshow test, p = 0.92). Negative and positive predictive values were 83.7% and 75%, respectively. The accuracy of the model was 82%. Therefore, assessment of interferon‐γ secretion by cytomegalovirus (CMV)‐specific CD8+ T cells prior to transplantation is useful in informing the risk of posttransplant CMV replication in solid organ transplant patients.     

Calcineurin/nuclear factor of activated T cells (NFAT) signaling in cobalt–chromium–molybdenum (CoCrMo) particles‐induced tumor necrosis factor‐α (TNFα) secretion in MLO‐Y4 osteocytes

Aseptic loosening is the devastating long term complication of total hip arthroplasty and orthopedic implant debris has been shown to trigger an intense inflammatory reaction leading to resorption of the bone matrix. Inflammatory cytokines, such as tumor necrosis factor‐α (TNFα), have been implicated in this process and osteocytes may play a role in its production. We previously demonstrated that cobalt–chromium–molybdenum (CoCrMo) particles upregulate TNFα production by MLO‐Y4 osteocytes in vitro, but the underlying mechanism has not been elucidated. Based on previous studies by others, we hypothesized that the calcineurin‐nuclear factor of activated T cells (NFAT) pathway mediates CoCrMo particle‐induced TNFα production in MLO‐Y4 osteocytes. MLO‐Y4 osteocytes exposed to CoCrMo particle treatment resulted in a rapid and significant increase in calcineurin activity. We also demonstrate that CoCrMo particle‐induced upregulation of TNFα is reduced to control levels with calcineurin‐NFAT inhibitors and this was also confirmed at mRNA level. Moreover, we demonstrate the localization of NFATs in MLO‐Y4 osteocytes and that NFAT1 and 2 translocate to the nucleus upon CoCrMo particle treatment. Our results suggest that calcineurin‐NFAT signaling is involved in TNFα production by MLO‐Y4 osteocytes after CoCrMo particle treatment. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:1867–1873, 2011     

直接削除法治疗鼻赘型酒糟鼻疗效

目的总结直接削除法治疗鼻赘型酒糟鼻的疗效。方法2015 年 1 月—2017 年 3 月，收治 6 例鼻赘型酒糟鼻男性患者。患者年龄 45～76 岁，平均 65 岁。病程 10～18 年，平均 15 年。术中以病变周边正常皮肤为参考平面削除赘生组织，并塑形鼻部各美学亚单位。术后创面隔天换药直至完全愈合。结果6 例创面顺利愈合。患者均获随访 6 个月。鼻部瘢痕无明显挛缩，鼻部各美学亚单位均保留，病变未复发。患者均对鼻部外形满意。结论直接削除法治疗鼻赘型酒糟鼻简单有效、易于掌握，术后鼻部外形较好，患者满意度高。     

Interleukin‐1β stimulates stromal‐derived factor‐1α expression in human subacromial bursa

Chemokines produced by synoviocytes of the subacromial bursa are up‐regulated in subacromial bursitis and rotator cuff disease. We hypothesized that SDF‐1α production in bursal synoviocytes may be induced by local cytokines such as interleukin IL‐1β and IL‐6. Subacromial bursa specimens were obtained from patients undergoing shoulder surgery. Bursal specimens were stained with anti‐human antibodies to IL‐1, IL‐6, and SDF‐1α by immunohistochemistry and compared to normal and rheumatoid controls. Bursal cells were also isolated from specimens and cultured. Early passaged cells were then treated with cytokines (IL‐1β and IL‐6) and SDF‐1α expression was measured by ELISA and RT‐PCR. SDF‐1α, IL‐1β, and IL‐6 were expressed at high levels in bursitis specimens from human subacromial bursa compared to normal controls. In cultured bursal synoviocytes, there was a dose‐dependent increase in SDF‐1α production in the supernatants of cells treated with IL‐1β. SDF‐1α mRNA expression was also increased in bursal cells treated with IL‐1β. IL‐6 caused a minimal but not statistically significant increase in SDF‐1α expression. SDF‐1α, IL‐1β, and IL‐6 are expressed in the inflamed human subacromial bursal tissues in patients with subacromial bursitis. In cultured bursal synoviocytes, SDF‐1α gene expression and protein production are stimulated by IL‐1β. IL‐1β produced by bursal syvoviocytes and inflammatory cells in the human subacromial bursa is an important signal in the inflammatory response that occurs in subacromial bursitis and rotator cuff disease. © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29:1695–1699, 2011     

Combined effects of TNF‐α, IL‐1β, and HIF‐1α on MMP‐2 production in ACL fibroblasts under mechanical stretch: An in vitro study

The dynamics between inflammatory factors, mechanical stress, and healing factors, in an intra‐articular joint, are very complex after injury. Injury to intra‐articular tissue [anterior cruciate ligament (ACL), synovium] results in hypoxia, accumulation of various pro‐inflammatory factors, cytokines, and metalloproteases. Although the presence of increased amounts of matrix‐metalloproteinases (MMP) in the joint fluid after knee injury is considered the key factor for ACL poor healing ability; however, the exact role of collective participants of the joint fluid on MMP‐2 activity and production has not been fully studied yet. To investigate the combined effects of mechanical injury, inflammation and hypoxia induced factor‐1α (HIF‐1α) on induction of MMP‐2; we mimicked the microenvironment of joint cavity after ACL injury. The results show that TNF‐α and IL‐1β elevate the activity of MMP‐2 in a dose‐ and time‐dependent manner. In addition, mechanical stretch further enhances the MMP‐2 protein levels with TNF‐α, IL‐1β, and their mixture. CoCl₂‐induced HIF‐1α (100 and 500 µM) also increases the levels and activity of MMP‐2. Mechanical stretch has a strong additional effect on MMP‐2 production with HIF‐1α. Our results conclude that mechanical injury, HIF‐1α and inflammatory factors collectively induce increased MMP‐2 production in ACL fibroblasts, which was inhibited by NF‐κB pathway inhibitor (Bay‐11‐7082). © 2011 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 29: 1008–1014, 2011     

Association between interleukin‐4R and TGF‐β1 gene polymorphisms and the risk of colorectal cancer in a Korean population

Aim Colorectal cancer is associated with inflammatory bowel disease. The mechanisms of how different genetic make‐ups of cytokines might influence the individual susceptibility to develop particular types of tumours are still unknown. The authors analysed the association between genetic polymorphisms in cytokine/cytokine receptor genes and the risk of colorectal cancer in a Korean population. Method The authors assessed polymorphisms of the interleukin: IL‐1, IL‐1R, IL‐2, IL‐4, IL‐4R, IL‐10, transforming growth factor (TGF)‐β1, IFN‐γ genes in Korean patients with colorectal cancer (n = 170) and in a normal healthy control group (n = 130) to investigate the association between theses cytokine gene polymorphisms and the risk of colorectal cancer. Results The IL‐4R 1902*T allele was found to be associated with an increased risk of colon cancer (P < 0.01, OR = 2.0) and rectal cancer (P < 0.05, OR = 1.8). The IL‐4R 1902*C allele was associated with a decreased risk of both colon cancer (P < 0.01, OR = 0.51) and rectal cancer (P < 0.05, OR = 0.5). The TFG‐β1 10*T allele was found to be associated with an increased risk of colon cancer (P < 0.00, OR = 2.3) and the TFG‐β1 10*C allele with a decreased risk of colon cancer (P < 0.00, OR = 0.43). Conclusion These results suggest that the genetic polymorphisms of IL‐4R and TGF‐β1 are associated with the risk of colorectal cancer in a Korean population.     

臀部筋膜脂肪瓣修复坐骨结节和大转子复发性窦道型压疮

目的总结臀部筋膜脂肪瓣修复坐骨结节、大转子复发性窦道型压疮的效果。方法2018 年 2 月—2019 年 6 月，收治 12 例 13 处长期截瘫伴坐骨结节、大转子复发性窦道型压疮患者。其中男 10 例 11 处，女 2 例 2 处；年龄 46～56 岁，平均 51 岁。截瘫 10～20 年，平均 13 年；所有患者均有压疮手术史，术后 3 个月～12 年复发。其中坐骨结节处压疮 11 例，坐骨结节合并大转子处压疮 1 例。创面清创、切除窦道假性滑液囊，采用单侧或双侧臀部筋膜脂肪瓣填塞窦道，术区一期缝合闭合切口。结果术后 13 处压疮切口均Ⅰ期愈合，局部无红肿、渗液，术后 14 d 拆线出院。术后局部平坦，外观理想。术后患者均获随访，随访时间 8～24 个月，平均 14 个月。随访期间压疮均无复发。结论臀部脂肪组织丰富，利用筋膜脂肪瓣修复坐骨结节、大转子复发性窦道型压疮设计、操作简便，临床效果良好。