Preservation of Residual Limb Length with Antibiotic-loaded Bone Cement Implantation to Treat Femoral Periprosthetic Infection: A Case Report.

BACKGROUND: : Fitting a femoral prosthesis in a transfemoral amputee with a very short amputation stump is challenging. This case report aimed to introduce an effective and simple method that can preserve the residual limb length by the implantation of antibiotic-loaded bone cement for the treatment of a patient with femoral periprosthetic infection. CASE: : A 30-year-old man who had osteosarcoma at the age of 13 years underwent transfemoral amputation 17 years after the initial surgery because of periprosthetic infection. Antibiotic-loaded bone cement was inserted into the infected bone marrow to control the residual infection and to preserve the stump length. The infection resolved, and the patient regained functional gait using a femoral prosthesis. DISCUSSION: : This case report demonstrates the usefulness of antibiotic-loaded cement in preserving the length of residual limbs and for femoral prosthesis fitting after periprosthetic infection. Maintaining the residual bone length is crucial in amputees for the functional fitting of femoral prostheses. The use of antibiotic-loaded bone cement has potential as a simple and useful surgical option in amputees after periprosthetic infection.

A case of intra-articular fasciitis in the elbow joint.

INTRODUCTION: Nodular fasciitis is a benign myofibroblastic proliferation arising from the fascia. Until now, there have been only two reported cases of intra-articular nodular fasciitis in the elbow joint. PRESENTATION OF CASE: We report a case of a 19-year-old woman with a 3-month history of pain in the left elbow. Contrast-enhanced T1-weighted magnetic resonance imaging (MRI) showed an intra-articular lobulated mass on the anterior portion of the elbow joint, with accompanying effusion. The patient subsequently underwent arthroscopic excision of the mass. Histologically, intra-articular nodular fasciitis was the final diagnosis. At the most recent follow-up, 20 months after surgery, the patient had no subjective symptoms, including pain. The final MRI findings showed no tumor recurrence. DISCUSSION: As nodular fasciitis is not generally known to arise within a joint, the occurrence at such anatomical locations may lead to a misdiagnosis. Intra-articular nodular fasciitis is rarely encountered, and therefore, is not usually considered during the clinical investigation of joint symptoms. CONCLUSION: Preoperative diagnosis was difficult in this case because of nonspecific preoperative clinical findings. Although histological examination is necessary to establish a diagnosis, we recommend that intra-articular nodular fasciitis should be included in the differential diagnosis of intra-articular mass lesions.

Chloroquine Enhances Rapamycin-induced Apoptosis in MG63 Cells.

BACKGROUND/AIM: We previously showed that the use of autophagy inhibitors in combination with chemotherapy can enhance anticancer effects in sarcoma cell lines. In this study, we investigated the combined effect of the autophagy inhibitor chloroquine and the mTOR inhibitor rapamycin on MG63 osteosarcoma cells. MATERIALS AND METHODS: Effects of chloroquine and/or rapamycin on cell proliferation were assessed by WST-1 assays. Effects of chloroquine and/or rapamycin on the mTOR pathway components, autophagy, and apoptosis were investigated by western blot, flow cytometry, and fluorescence microscopy using immunocytochemical staining of LC3 and Annexin V-FITC/propidium iodide. RESULTS: Rapamycin suppressed cell growth and inhibited the mTOR pathway. Rapamycin promoted autophagy by blocking the mTOR pathway, and chloroquine enhanced apoptosis by blocking autophagy. CONCLUSION: Chloroquine enhances the effects of rapamycin in inducing apoptosis via autophagy inhibition in MG63 cells. Thus, the combined therapy of chloroquine and rapamycin may be a potent treatment for osteosarcoma.

SNX-2112 Induces Apoptosis and Autophagy of Nara-H Cells.

BACKGROUND/AIM: Selective heat shock protein 90 (Hsp90) inhibitor SNX-2112 exhibits antitumor activity in multiple cancer cell types. Here, the antitumor activity of SNX-2112 in Nara-H cells was analyzed. MATERIALS AND METHODS: Antitumor activity of SNX-2112 was assessed using a cell proliferation assay. We also examined the signalling pathways involved in SNX-2112-mediated autophagy and apoptosis of Nara-H cells by western blot and morphological analyses. RESULTS: Cell proliferation assays demonstrated that SNX-2112 inhibited Nara-H cell growth. Western blotting revealed that SNX-2112 induced apoptosis and autophagy, inhibited mammalian target of rapamycin (mTOR) phosphorylation, and suppressed the mitogen-activated protein kinase (MAPK) signalling pathway. Morphological analysis confirmed that SNX-2112 induced autophagy and apoptosis. CONCLUSION: SNX-2112 induced autophagy and apoptosis of Nara-H cells by inhibiting mTOR and MAPK pathways. Our results support developing SNX-2112 to treat human soft tissue sarcomas.

A rare case of acute osteomyelitis due to Panton-Valentine leukocidin-positive community-acquired methicillin-resistant Staphylococcus aureus in a young healthy adult.

INTRODUCTION: Most community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) infections affect skin or soft tissues, while invasive and life-threatening illnesses including osteomyelitis are less common. CA-MRSA infections occur especially in the pediatric age group, while the occurrence of CA-MRSA osteomyelitis in adults is uncommonly reported. PRESENTATION OF CASES: A rare case of acute osteomyelitis of the femur caused by Panton-Valentine leukocidin (PVL)-positive CA-MRSA in a 37-year-old man in good health is presented. A pure bone biopsy revealed extensive inflammation, suggestive of acute osteomyelitis, with no evidence of neoplasm, and PVL-positive MRSA was isolated from the culture. Antibiotic treatment, with 6 weeks of intravenous vancomycin and 4 weeks of clindamycin, followed by 2 weeks of oral linezolid, was given, and 2 years after treatment completion, there has been no relapse of infection. CONCLUSION: This case strongly suggests that we need to be aware of CA-MRSA osteomyelitis, which requires a high level of suspicion, prompt diagnosis, and appropriate antibiotic treatment.

Effects of suppressed bone remodeling by minodronic acid and alendronate on bone mass, microdamage accumulation, collagen crosslinks and bone mechanical properties in the lumbar vertebra of ovariectomized cynomolgus monkeys.

Collagen crosslinking is an important determinant of the quality of bone material. We have previously shown that suppressed bone turnover by high doses of minodronic acid and alendronate increases compressive strength of vertebra, but also increases microdamage accumulation, in monkey bone. The aim of this study is to examine the effects of these bisphosphonates on collagen crosslinks and intrinsic material properties, in addition to microdamage accumulation, in vertebral cancellous bone in ovariectomized cynomolgus monkeys. Sixty female monkeys aged 9-17years were divided into five groups: sham and ovariectomized groups were treated daily for 17months with lactose vehicle, and the other three groups were given minodronic acid daily at 0.015 or 0.15mg/kg or alendronate daily at 0.5mg/kg orally. After sacrifice, lumbar vertebrae were subjected to histomorphometry, microdamage measurement, analysis of collagen crosslinking and compressive mechanical tests. Minodronic acid caused dose-dependent suppression of increased bone remodeling due to ovariectomy, and low-dose minodronic acid suppressed remodeling same level as alendronate. However, low-dose minodronic acid did not change microdamage accumulation, collagen maturity and the pentosidine level, whereas high-dose minodronic acid and alendronate increased these parameters. Compressive ultimate load was increased following high-dose minodronic acid and alendronate, but no treatment altered the reduction in intrinsic material properties caused by ovariectomy. These findings suggest that deterioration of bone material and formation of pentosidine and microdamage induced by minodronic acid is less than that expected based on the extent of remodeling suppression, in comparison with alendronate, but this was not reflected in any significant change of mechanical properties.

GSK-3 inhibitor inhibits cell proliferation and induces apoptosis in human osteosarcoma cells.

Glycogen synthase kinase-3 (GSK-3) is a serine/threonine kinase that functions in numerous signaling pathways initiated by diverse stimuli. The functions of GSK-3 in cancer differ depending on cell type. In the present study, we examined the effects of a specific GSK-3 inhibitor on the regulation of osteosarcoma cell proliferation and apoptosis. Immunohistochemical analysis and real-time reverse transcription-polymerase chain reaction (RT-PCR) were performed to determine the expression pattern of GSK-3 in human osteosarcoma cells. We used the MTS assay, western blotting, measurement of single-stranded DNA and morphological analyses to study the effects of a GSK-3 inhibitor, SB216763 on osteosarcoma cell proliferation and survival. We detected an increase in mRNA expression of GSK-3 and aberrant nuclear accumulation of GSK-3 in the osteosarcoma cells. Pharmacological inhibition of GSK-3 led to a decrease in proliferation and survival of osteosarcoma cells. Inhibition of GSK-3 resulted in a decreased expression of Bcl-2 and a subsequent increase in osteosarcoma cell apoptosis via the mitochondrial pathway. The present study demonstrated that GSK-3 activity is critical for tumorigenicity and cell survival in osteosarcoma cells. Our findings suggest that GSK-3 is a potential therapeutic target for the treatment of human osteosarcoma.

Apoptosis and antitumor effects induced by the combination of an mTOR inhibitor and an autophagy inhibitor in human osteosarcoma MG63 cells.

The inhibition of the mammalian target of rapamycin (mTOR) signaling pathway promotes the initiation of autophagy. Although it remains under debate whether chemotherapy-induced autophagy in tumor cells is a protective response or is invoked to promote cell death, recent studies indicate that autophagy is a self-defense mechanism of cancer cells that are subjected to antitumor agents and that blocking autophagy can trigger apoptosis. The aim of this study was to examine the effects of rapamycin, an mTOR inhibitor, on MG63 osteosarcoma cells. We further examined whether the combination of rapamycin and the small molecule inhibitor of autophagy Spautin-1 (specific and potent autophagy inhibitor-1) enhanced the rapamycin-induced apoptosis in MG63 cells. We examined the effects of rapamycin treatment on cell proliferation, phosphorylation of mTOR pathway components, and autophagy by western blot analysis. Furthermore, we examined the effects of rapamycin with or without Spautin-1 on the induction of apoptosis by western blot analysis and immunohistochemical staining. We found that rapamycin inhibited cell proliferation and decreased the phosphorylation of mTOR pathway components in MG63 cells. Rapamycin induced the apoptosis of MG63 cells, and this apoptosis was enhanced by Spautin-1. It was considered that Spautin-1 suppressed the protective mechanism induced by rapamycin in tumor cells and induced apoptosis. Therefore, the combination of an mTOR inhibitor and an autophagy inhibitor may be effective in the treatment of osteosarcoma because it effectively induces the apoptotic pathway.

A Rare Case of Tumoral Calcium Pyrophosphate Dihydrate Crystal Deposition Disease of the Wrist Joint.

Introduction. Tumoral calcium pyrophosphate dihydrate (CPPD) crystal deposition disease (CPPDCD), also known as tophaceous calcium pyrophosphate deposition disease (CPDD), is a tumorlike lesion, and it should be distinguished from usual CPDD that causes severe joint inflammation and arthralgia. A case of tumoral CPPDCD of the wrist joint that required differentiation from synovial osteochondromatosis is described. Case Presentation. The patient was a 78-year-old woman with a 5-year history of nodular lesions at the right wrist that had gradually increased in size. An excisional biopsy and a histological examination of the excised nodular lesions by hematoxylin and eosin (H&E) staining were performed, demonstrating numerous polarizable, rhabdoid, and rectangular crystals, surrounded by fibroblasts, macrophages, and foreign body-type giant cells, consistent with tumoral CPPDCD. Conclusion. Tumoral CPPDCD, especially at the wrist joint, is rare, and, to the best of our knowledge, only 2 articles have been published. This case seems to need further follow-up for recurrence, because tumoral CPPDCD may recur after complete or incomplete surgical excision.

Hsp90 inhibitor induces autophagy and apoptosis in osteosarcoma cells.

Heat shock protein 90 (Hsp90) is constitutively expressed at 2­10­fold higher levels in tumor cells compared to normal cells, suggesting that it may be critically important for tumor cell growth and survival. These features make Hsp90 a potential target for anticancer drug development. Inhibition of Hsp90 activity not only results in rapid degradation of Hsp90 client proteins but also induces apoptosis of various tumor cells. Hsp90 also plays an important role in autophagy. An Hsp90 inhibitor induces autophagy through inhibition of mTOR. It is still under debate whether chemotherapy­induced autophagy in tumor cells is a protective response or is invoked to promote cell death. The aim of this study was to examine the effects of the Hsp90 inhibitor, geldanamycin (GA), on KTHOS osteosarcoma cells. We further examined whether a combination of GA and the autophagy inhibitor 3­methyladenine (3­MA) enhanced GA­induced apoptosis in KTHOS cells. GA had an inhibitory effect on cell proliferation and inhibited the Akt/mTOR signaling pathway in KTHOS cells. GA alone induced autophagy and apoptosis in KTHOS cells, but treatment with a combination of GA and 3­MA suppressed autophagy and induced apoptosis to a much greater extent than GA alone in these cells. It was considered that the autophagy inhibitor 3­MA suppressed a protective mechanism induced by Hsp90 inhibitor in tumor cells and induced apoptosis. Therefore, the combination of an Hsp90 inhibitor and an autophagy inhibitor may be an effective treatment for osteosarcoma because this combination effectively induces apoptotic pathways.

A large amount of microdamages in the cortical bone around fracture site in a patient of atypical femoral fracture after long-term bisphosphonate therapy.

A breast cancer patient developed an atypical femoral fracture after 9 years of bisphosphonate therapy for the treatment of multiple bone metastases. We histopathologically analyzed the femoral cortical bone at the fracture site and the iliac cancellous bone. Four months prior to the fracture, the patient had experienced pain in the right femur and underwent plain radiography and bone scintigraphy which revealed cortical thickening and radioisotope accumulation at each site, respectively. The patient had also experienced a non-traumatic fracture at the same site on the contralateral side 2 years earlier. Based on these findings, atypical femoral fracture was diagnosed and intramedullary nailing performed. A cortical bone specimen taken from near the fracture site during surgery showed marked microdamages, and analysis of the iliac cancellous bone specimen revealed severely suppressed bone turnover. These findings suggest that microdamage and severely suppressed bone turnover are associated with atypical femoral fracture reported in this patient with long-term bisphosphonate therapy.

The combination of rapamycin and MAPK inhibitors enhances the growth inhibitory effect on Nara-H cells.

The inhibition of the mammalian target of rapamycin (mTOR) signaling pathway promotes the initiation of autophagy, and the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated protein kinase (ERK) is well known to induce autophagy. Autophagy is a self-defense mechanism of cancer cells that are subjected to antitumor agents, and blocking autophagy can trigger apoptosis. In the present study, we demonstrate that an mTOR inhibitor, rapamycin, induces autophagy in the Nara-H malignant fibrous histiocytoma (MFH) cell line through the activation of ERK1/2. Rapamycin-induced apoptosis was enhanced following the inhibition of the MEK/ERK pathway. In the Nara-H cells, we examined the effects of rapamycin treatment on cell proliferation and on the phosphorylation of the mTOR pathway components and autophagy by western blot analysis. Furthermore, we examined the effects of rapamycin with or without the MEK inhibitor, U0126, on the induction of apoptosis by using fluorescence microscopy. Rapamycin inhibited Nara-H cell proliferation and decreased the phosphorylation of the mTOR pathway in the Nara-H cells. Rapamycin induced the apoptosis of Nara-H cells, and this apoptosis was enhanced by U0126. Simultaneously, phospho-ERK1/2 was activated by rapamycin. The present study demonstrates that rapamycin induces autophagy in Nara-H cells by activating the MEK/ERK signaling pathway, and the rapamycin-induced apoptosis can be enhanced by the MEK inhibitor, U0126. These results suggest that self­protective mechanisms involving mTOR inhibitors in Nara-H cells are prevented by the inhibition of the MEK/ERK pathway. The combination of an mTOR inhibitor (e.g., rapamycin) and an MEK inhibitor (e.g., U0126) may offer effective treatment for MFH, as this combination effectively activates apoptotic pathways.

Polyostotic fibrous dysplasia with epiphyseal involvement in long bones: a case report.

Fibrous dysplasia (FD) is an uncommon, but well-known benign skeletal disorder. In cases affecting long bones, FD is commonly recognized to locate in the diaphyses or the metaphyses and to spare the epiphyses. In this paper, we present a rare case of polyostotic FD in a 13-year-old girl with unilateral multiple epiphyseal lesions arising in the femur, the tibia, and the fibula with the growth plates.

Prefabrication of vascularized bone allograft in a recipient rat using a flow-through vascular pedicle, bone morphogenetic protein, and bisphosphonate.

We attempted to prefabricate vascularized bone allografts by implanting flow-through vascular bundles from recipient rats into transplanted bone allografts. We also applied bone morphogenetic protein (BMP) and bisphosphonate into the bone allograft to accelerate bone formation and inhibit bone resorption in the transplanted bone. After prefabrication, bone formation and resorption in the vascularized bone allograft were evaluated radiographically and histologically. We also attempted to transfer the prefabricated vascularized bone allograft onto the femur of recipient rats, and bone union between was subsequently assessed. Bone formation in the transplanted allograft was significantly stimulated with addition of BMP. However, bone resorption was also stimulated by BMP; this stimulated bone resorption caused by BMP was effectively inhibited with addition of bisphosphonate. The bone union rate between transplanted bone allografts and recipient femora was also stimulated by BMP. Bisphosphonate slightly delayed bone union but effectively protected the grafted bone from bone resorption caused by BMP. Our results suggest that prefabrication of vascularized bone allografts can be achieved in the recipient rat by implanting a flow-through vascular bundle from the recipient into the transplanted bone allograft. Combination treatment with BMP and bisphosphonate allows development of an ideal vascularized bone allograft.

Effects of minodronic acid and alendronate on bone remodeling, microdamage accumulation, degree of mineralization and bone mechanical properties in ovariectomized cynomolgus monkeys.

Suppression of bone remodeling by bisphosphonates leads to accumulation of microdamage in bone. If this microdamage develops due to suppressed repair of remodeling only, more potent bisphosphonates should cause more damage. In this study, we evaluated the effects of reduced bone turnover produced by a potent bisphosphonate, minodronic acid, on microdamage accumulation, the degree of mineralization and mechanical properties of bone in ovariectomized cynomolgus monkeys, and compared these effects with those of alendronate. Sixty female monkeys aged 9-17 years old were divided into five groups. The sham group and the ovariectomized group were treated daily for 17 months with lactose vehicle. The other three groups were treated daily with minodronic acid at a dose of 0.015 mg/kg or 0.15 mg/kg, or alendronate at 0.5mg/kg orally. After sacrifice, lumbar vertebrae and left femurs were subjected to histomorphometry, microdamage, mineralization analyses, and mechanical testing. Minodronic acid suppressed bone remodeling of cancellous and cortical bone in a dose-dependent manner and the higher dose of minodronic acid suppressed bone remodeling more strongly than alendronate. The lower dose of minodronic acid did not increase microdamage accumulation and compressive strength, but the higher dose of minodronic acid and alendronate resulted in similar increases in cancellous microdamage accumulation and ultimate load in lumbar vertebra. There were no significant differences among the groups in microdamage, degree of mineralization and mechanical properties in cortical bone of the femoral shaft; however, only alendronate showed a tendency to increase highly mineralized osteons and microdamage. These findings suggest that microdamage caused by minodronic acid is less than that expected based on the extent of remodeling suppression, in comparison with alendronate although this was not reflected in any significant change of mechanical properties.

Massive heterotopic ossification around the ankle in a patient with macrodactyly of the foot: a case report.

A 40-year-old man with macrodactyly of the left great toe presented with progressive enlargement of the ipsilateral ankle and increasing pain after minor trauma. Radiographs and computed tomographic scans of the foot and ankle revealed macrodactyly of the great toe and a large heterotopic ossified mass in the anteromedial aspect of the ankle. The large osseous lesion in the ankle was operatively excised. The lesion was easily excised en bloc after releasing the adhesion and histologically diagnosed as normal mature bone with osteoarthritic changes. Massive heterotopic ossification developing around the ankle in a patient with macrodactyly has not been previously reported. The current case was classified as hyperostotic macrodactyly. Operative treatment relieved the pain and improved the range of the motion of the ankle.

Inhibition of induced autophagy increases apoptosis of Nara-H cells.

Inhibition of the mTOR signaling pathway promotes initiation of autophagy. However, recent studies indicate that autophagy is a self-defense mechanism of cancer cells that are subjected to anti-tumor agents and that blocking autophagy can trigger apoptosis. Here, we examined the effects of an mTOR inhibitor, temsirolimus, on a malignant fibrous histiocytoma (MFH) cell line, Nara-H cells, and the effect of suppressing autophagy on the induction of apoptosis in these MFH cells. In Nara-H cells, we examined the effects of temsirolimus treatment on cell proliferation using the CellTiter 96® AQueous One Solution Cell Proliferation Assay and on phosphorylation of mTOR pathway components and autophagy using Western blot-based assays. Furthermore, we examined the effects of temsirolimus with or without 3-methyladenine (3-MA) on induction of apoptosis using fluorescent microscopic analysis. In Nara-H cells, temsirolimus treatment inhibited cell proliferation, suppressed phosphorylation of mTOR pathway components, and induced autophagy as assessed by LC-3 II expression. Moreover, treatment with a combination of temsirolimus and 3-MA induced apoptosis in Nara-H cells. Apparently, simultaneous inhibition of autophagy and mTOR induced apoptosis in Nara-H cells because inhibition of autophagy prevented the cells from protecting themselves from the effects of the inhibition of mTOR. Therefore, a combination therapy that includes an mTOR inhibitor and an autophagy inhibitor (temsirolimus and 3-MA, respectively) may effectively treat MFH by inducing apoptosis in tumor cells.

Percutaneous curettage and continuous irrigation for MRSA lumbar spondylodiscitis: a report of three cases.

There has been a recent increase in pyogenic spondylitis caused by methicillin-resistant Staphylococcus aureus (MRSA) associated with an increasing number of compromised patients. As long as serious paralysis is absent, we recommend percutaneous curettage and continuous irrigation as an effective treatment for MRSA lumbar spondylodiscitis. Under local anesthesia, the affected lumbar discs were curetted using percutaneous nucleotomy, and tubes were placed for continuous irrigation. The period of continuous irrigation was generally 2 weeks. Infection was controlled after one procedure in two cases and after two procedures in one case. Postoperative radiography and magnetic resonance imaging (MRI) showed callus formation, normalized signal intensity in vertebral bodies, and regression of abscesses. Open surgery under general anesthesia has been considered risky in patients with poor performance status or old age. The present method, which is an application of needle biopsy, can be performed under local anesthesia and is minimally invasive.