Mesenchymal Stem Cells Preconditioned with Hypoxia and Dexamethasone Promote Osteoblast Differentiation Under Stress Conditions.

Bone marrow-derived mesenchymal stem cells (MSCs), which are capable of differentiating into osteoblasts, are used in effective regenerative therapies. MSCs must be prompted to differentiate into osteoblasts for MSC transplantation to be effective. In this study, osteoblast differentiation markers involved in bone formation were evaluated to investigate the stress resistance of bone marrow-derived rat MSCs to dexamethasone and hypoxia and their ability to differentiate into osteoblasts. MSCs were allowed to differentiate into osteoblasts for 21 days in three different environments (dexamethasone treatment, hypoxic conditions [1% oxygen], or both). Osteoblast differentiation potential was evaluated according to alkaline phosphatase levels and a mineralisation assay. Immunofluorescence staining was used to determine the protein expression of the osteoblast differentiation markers type I collagen and osteopontin. MSCs differentiated into osteoblasts under hypoxic conditions but differentiated more slowly upon treatment with dexamethasone and dexamethasone plus hypoxia relative to the control. MSCs preconditioned with dexamethasone or hypoxia and then allowed to differentiate into osteoblasts under similar conditions differentiated comparably to control MSCs. MSCs that developed resistance to dexamethasone or hypoxia differentiated more quickly into osteoblasts than those that did not. The findings suggest that increasing the resistance of MSCs to stress by preconditioning them via dexamethasone or hypoxia exposure could result in more rapid differentiation into osteoblasts following transplantation.

[A rare case of intraductal tumor of the pancreas in which an intraductal tubulopapillary neoplasm was mixed with a widely spreading gastric-type intraductal papillary-mucinous neoplasm].

A 70-year-old man receiving treatment for diabetes mellitus presented with a cystic mass in the border area of the pancreatic body and tail on plain computed tomography (CT) due to impaired glucose intolerance. Contrast-enhanced CT showed a faint hyperattenuated nodular mass extending from the dilated main pancreatic duct (MPD) to the branch duct. Endoscopic retrograde cholangiopancreatography revealed a mildly dilated orifice of the papilla of Vater and MPD stenosis with entire upstream and immediate downstream dilatations. The patient underwent distal pancreatectomy due to the suspicion of mixed-type intraductal papillary-mucinous carcinoma. A pathological examination showed an intraductal solid-nodular mass measuring 25mm in length, consisting of two types of neoplasms. One showed tubulopapillary growth with entirely high-grade (HG) atypical cuboidal epithelium, in which immunohistochemical examinations were positive for MUC6 but negative for human gastric mucin (HGM), MUC1, MUC2, and MUC5AC, fitting the concept of intraductal tubulopapillary neoplasm (ITPN). The other showed the same growth of low-grade (LG) atypical columnar cells positive for HGM and MUC5AC and negative for MUC1 and MUC2, which corresponded to gastric-type intraductal papillary-mucinous neoplasm (IPMN) -LG. The tumor had not invaded the duct walls, and no metastatic lymph nodes were observed. The ITPN was adjacent to the IPMN mainly composed of tubular glands mimicking pyloric glands with LG dysplasia that corresponded to the so-called IPMN-pyloric gland variant. Moreover, the proliferation of low-papillary gastric-type IPMN spread around the intraductal tumors. Consequently, the patient was diagnosed with an intraductal tubular neoplasm comprising a noninvasive ITPN and gastric-type IPMN-LG. ITPN is a recently identified intraductal neoplasm of the pancreas proposed by Yamaguchi et al. and is distinguished by intraductal tubulopapillary growth with HG cellular atypia without overt mucin production, in contrast to IPMN. To date, no cases of intraductal nodular tumors comprising ITPN and IPMN have been reported. We report this original case with imaging and pathological observations and discuss potential processes via which ITPN and IPMN may arise adjacent to each other in the same pancreatic duct.

[Carcinoma in situ arising in adenomyomatous hyperplasia of the ampulla of Vater: the first case report].

A man in his 70s was admitted to our hospital due to jaundice and upper abdominal pain. Laboratory findings indicated elevated serum hepatobiliary enzyme and amylase levels. Contrast-enhanced computed tomography revealed smooth wall thickening of the terminal bile duct (tBD) with a faintly enhanced inner line. ERCP revealed stenosis from the tBD to the ampulla of Vater (AV) with upstream dilatation. Intraductal ultrasound (IDUS) circumferentially revealed a thickened wall preserving a three-layered structure throughout the same region. Furthermore, a thick innermost hyperechoic layer was identified in the bile duct portion of the AV (Ab). Findings suggestive of adenocarcinoma were obtained from the tissue samples from the biliary stricture using biopsy forceps. Thus, pancreatoduodenectomy was performed. A pathological examination revealed a thickened AV wall spreading over the tBD with hyperplasia of the glands and smooth muscle fibers. In addition, low-grade biliary intraepithelial neoplasia (BilIN) was scattered throughout the lesion, and high-grade BilIN was partly observed in the peribiliary glands of the Ab. Based on these results, a diagnosis of carcinoma in situ arising in adenomyomatous hyperplasia (ADMH) of the AV was made. To date, there are no reports on ADMH-associated carcinoma of the BD or AV. We here report this original case with the IDUS findings, which are presumed to reflect the histologic features of ADMH showing ductal proliferation surrounded by smooth muscle fibers. Also, we discuss the process through which carcinoma arises from ADMH in AV.

Surgical implantation of human adipose derived stem cells attenuates experimentally induced hepatic fibrosis in rats.

BACKGROUND: Mesenchymal stem cells (MSCs) are multipotent stromal cells and could exert hepatoprotective effects against acute liver injury, steatohepatitis, and fibrogenesis. Here, we evaluated the effects of human adipose derived stem cells (hADSCs) to attenuate experimentally induced hepatic fibrosis and early cirrhosis in rats. METHODS: Hepatic fibrosis was induced by intraperitoneal injections of CCl4 (0.1 ml/100 g body weight) twice a week for 8 weeks. hADSCs were isolated and cultured on polyethylene discs coated with hydroxyapatite and 2 cm diameter disc was surgically implanted on the right lateral lobe of the liver. Discs implanted without hADSCs served as control. The animals were injected again with CCl4 once a week for another 8 weeks. All the animals were sacrificed at the end of 16th week. RESULTS: Serial administrations of CCl4 resulted in well developed fibrosis and early cirrhosis at 8th week which maintained until the 16th week. Animals treated with hADSC discs depicted over 50% decrease of collagen with significant increase in serum albumin and total protein levels. Immunohistochemical staining for TGF-ß1, α-smooth muscle actin, and collagen type I and type III demonstrated marked decrease compared to the animals without hADSC treatment. CONCLUSIONS: Treatment with hADSCs improved liver functions, markedly reduced hepatic fibrosis and early cirrhosis. Various pleiotropic and paracrine factors secreted from the hADSCs seem to serve as reparative functions in the attenuation of liver cirrhosis. The data demonstrated that treatment with hADSCs can be successfully used as a potent therapeutic method to prevent progression of hepatic fibrosis and related adverse events.

Resistance of bone marrow mesenchymal stem cells in a stressed environment - Comparison with osteocyte cells.

Introduction: Recently, the efficacy of mesenchymal stem cells (MSCs) mediated by their tissue repair and anti-inflammatory actions in the prevention and therapy of various disorders has been reported. In this research, our attention was focused specifically on the prevention and therapy of glucocorticoid-induced osteonecrosis. We investigated the stress resistance of MSC against glucocorticoid administration and hypoxic stress, which are factors known to induce osteocytic cell death. Materials and Methods: Mouse bone cells (MLO-Y4) and bone-marrow derived mouse MSCs were exposed to dexamethasone (Dex), hypoxia of 1% oxygen or both in vitro. Mitochondrial membrane potentials were estimated by mitochondria labeling with a cell-permeant probe (Mito tracker red); expression of these apoptosis-inducing molecules, oxidative stress marker (8-hydroxy-2'-deoxyguanosine), caspase-3, -9, and two apoptosis-inhibiting molecules, energy-producing ATP synthase (ATP5A) and X-linked inhibitor of apoptosis protein (XIAP), were analyzed by both immunofluorescence and western blot. Results: With exposure to either dexamethasone or hypoxia, MLO-Y4 showed reduced mitochondrial membrane potential, ATP5A and upregulation of 8-OHdG, cleaved caspases and XIAP. Those changes were significantly enhanced by treatment with dexamethasone plus hypoxia. In MSCs, however, mitochondrial membrane potentials were preserved, while no significant changes in the pro-apoptosis or anti-apoptosis molecules analyzed were found even with exposure to both dexamethasone and hypoxia. No such effects induced by treatment with dexamethasone, hypoxia, or both were demonstrated in MSCs at all. Discussion: In osteocyte cells subjected to the double stresses of glucocorticoid administration and a hypoxic environment osteocytic cell death was mediated via mitochondria. In contrast, MSC subjected to the same stressors showed preservation of mitochondrial function and reduced oxidative stress. Accordingly, even under conditions sufficiently stressful to cause the osteocytic cell death in vivo, it was thought that the function of MSC could be preserved, suggesting that in the case of osteonecrosis preventative and therapeutic strategies incorporating their intraosseous implantation may be promising.

Mitochondrial Transcription Factor A added to Osteocytes in a Stressed Environment has a Cytoprotective Effect.

The main precipitant of glucocorticoid-associated femoral head osteonecrosis is widely accepted to be an ischemic-hypoxic event, with oxidative stress also as an underlying factor. Mitochondrial DNA is more vulnerable to oxidative injury than the nucleus, and mitochondrial transcription factor A (TFAM), which plays roles in its function, preservation, and regulation is being increasingly investigated. In the present study we focused on the impact of TFAM on the relation between the oxidative injury induced by the addition of glucocorticoid to a hypoxic environment and osteocytic cell necrosis. Using cultured osteocytes MLO-Y4 in a 1% hypoxic environment (hypoxia) to which 1µM dexamethasone (Dex) was added (Dex(+)/hypoxia(+)), an immunocytochemical study was conducted using 8-hydroxy-2'-deoxyguanosine (8-OHdG), an index of oxidative stress, and hypoxia inducible factor-1α (HIF-1α), a marker of hypoxia. Next, after adding TFAM siRNA, TFAM knockdown, cultured for 24h, and mitochondrial membrane potential were measured, they were stained with ATP5A which labels adenosine triphosphate (ATP) production. Dex was added to MLO-Y4 to which TFAM had been added, and cultured for 24h in hypoxia. The ratio of dead cells to viable cells was determined and compared. Enhanced expression of 8-OHdG, HIF-1α was found in osteocytes following the addition of glucocorticoid in a hypoxic environment. With TFAM knockdown, as compared to normoxia, mitochondrial function significantly decreased. On the other hand, by adding TFAM, the incidence of osteocytic cell necrosis was significantly decreased as compared with Dex(+)/hypoxia(+). TFAM was confirmed to be important in mitochondrial function and preservation, inhibition of oxidative injury and maintenance of ATP production. Moreover, prevention of mitochondrial injury can best be achieved by decreasing the development of osteocytic cell necrosis.

Taurine Inhibits Glucocorticoid-Induced Bone Mitochondrial Injury, Preventing Osteonecrosis in Rabbits and Cultured Osteocytes.

Mitochondrial injury has recently been implicated in the pathogenesis of glucocorticoid-induced osteonecrosis. Using cultured osteocytes and a rabbit model, we investigated the possibility that taurine (TAU), which is known to play a role in the preservation of mitochondrial function, might also prevent the development of osteonecrosis. To reduplicate the intraosseous environment seen in glucocorticoid-induced osteonecrosis, dexamethasone (Dex) was added to MLO-Y4 cultured in 1% hypoxia (H-D stress environment). An in vitro study was conducted in which changes in mitochondrial transcription factor A (TFAM), a marker of mitochondrial function, and ATP5A produced by mitochondria, induced by the presence/absence of taurine addition were measured. To confirm the effect of taurine in vivo, 15 Japanese White rabbits were administered methylprednisolone (MP) 20 mg/kg as a single injection into the gluteus muscle (MP+/TAU- group), while for 5 consecutive days from the day of MP administration, taurine 100 mg/kg was administered to 15 animals (MP+/TAU+ group). As a control 15 untreated rabbits were also studied. The rabbits in each of the groups were sacrificed on the 14th day after glucocorticoid administration, and the bilateral femora were harvested. Histopathologically, the incidence of osteonecrosis was quantified immunohistochemically by quantifying TFAM and ATP5A expression. In the rabbits exposed to an H-D stress environment and in MP+/TAU- group, TFAM and ATP5A expression markedly decreased. With addition of taurine in the in vitro and in vivo studies, the expression of TFAM and ATP5A was somewhat decreased as compared with Dex-/hypoxia- or MP-/TAU- group, while improvement was noted as compared with Dex+/hypoxia+ or MP+/TAU- group. In rabbits, the incidence of osteonecrosis was 80% in MP+/TAU- group, in contrast to 20% in the taurine administered group (MP+/TAU+), representing a significant decrease. Since taurine was documented to exert a protective effect on mitochondrial function by inhibiting the mitochondrial dysfunction associated with glucocorticoid administration, we speculated that it might also indirectly help to prevent the development of osteonecrosis in this context. Since taurine is already being used clinically, we considered that its clinical application would also likely be smooth.

The combination of fatty liver and increased gamma-glutamyl transpeptidase levels as a risk factor for atherosclerotic plaque development in apparently healthy people

Background/aim: To elucidate how the combination of fatty liver and increased serum gamma-glutamyl transpeptidase (GGT) levels influences atherosclerotic plaque development in apparently healthy people. Materials and methods: The study population included people who had received an annual health checkup for more than 7 years and had no evidence of carotid plaque at baseline. We investigated the risk factors for carotid plaque occurrence using the Cox proportional hazards model. Results: A total of 107 people (76 men and 31 women; median age, 49 years) were enrolled. At baseline, fatty liver and a serum GGT level ≥50 U/L were observed in 13 and 38 people, respectively. During a median follow-up period of 13.3 years, carotid plaques appeared in 34 people. Multivariate analysis revealed that the combination of fatty liver and a serum GGT level ≥50 U/L was the only significant risk factor for carotid plaque occurrence (age- and sex-adjusted hazard ratio: 5.55; 95% confidence interval 1.70­18.14; P = 0.005). Conclusion: The combination of fatty liver and increased serum GGT levels raises the risk for atherosclerotic plaque development in apparently healthy people.

Mitochondrial stress and redox failure in steroid-associated osteonecrosis.

The purpose of the role of antioxidant enzymes and mitochondria in the developmental mechanism of steroid-associated osteonecrosis in the femur. In the present study Japanese white rabbits (mean weight 3.5kg) were injected into the gluteus with methylprednisolone (MP) 20mg/kg, and killed after 3 days (MP3 group), 5 days (MP5 group), and 14 days (MP14 group) (n=3 each). As a Control group (C group) Japanese white rabbits not administered MP were used. In experiment 1, the expression of the antioxidant enzymes Superoxide dismutade (SOD) and catalase was compared in liver, kidney, heart, humerus, and femur in C group, and the presence/absence of mitochondria transcription factor A (TFAM) expression was determined by Western blotting (WB) and used to evaluate the number of mitochondria and their function. In experiment 2, the presence/absence of necrosis was determined by immunohistochemistry, while changes in the expression of SOD, catalase, and TFAM in the femur after steroid administration were determined by Western blotting (WB). In experiment 1, intense expression of all of SOD, catalase, and TFAM was found in the liver, kidney, and heart as compared to the humerus and femur. In experiment 2, the expression of all of SOD, catalase, and TFAM in MP3 and MP5 groups was decreased on WB as compared with C group, while in MP14 group a tendency to improvement was seen. Accordingly, steroid-associated mitochondrial injury and redox failure are concluded to be important elements implicated in the pathogenesis of osteonecrosis.

Angiogenesis after administration of basic fibroblast growth factor induces proliferation and differentiation of mesenchymal stem cells in elastic perichondrium in an in vivo model: mini review of three sequential republication-abridged reports.

To date, studies on mesenchymal tissue stem cells (MSCs) in the perichondrium have focused on in vitro analysis, and the dynamics of cartilage regeneration from the perichondrium in vivo remain largely unknown. We have attempted to apply cell and tissue engineering methodology for ear reconstruction using cultured chondrocytes. We hypothesized that by inducing angiogenesis with basic fibroblast growth factor (bFGF), MSCs or cartilage precursor cells would proliferate and differentiate into cartilage in vivo and that the regenerated cartilage would maintain its morphology over an extended period. As a result of a single administration of bFGF to the perichondrium, cartilage tissue formed and proliferated while maintaining its morphology for at least 3 months. By day 3 post bFGF treatment, inflammatory cells, primarily comprising mononuclear cells, migrated to the perichondrial region, and the proliferation of matrix metalloproteinase 1 positive cells peaked. During week 1, the perichondrium thickened and proliferation of vascular endothelial cells was noted, along with an increase in the number of CD44-positive and CD90-positive cartilage MSCs/progenitor cells. Neocartilage was formed after 2 weeks, and hypertrophied mature cartilage was formed and maintained after 3 months. Proliferation of the perichondrium and cartilage was bFGF concentration-dependent and was inhibited by neutralizing antibodies. Angiogenesis induction by bFGF was blocked by the administration of an angiogenesis inhibitor, preventing perichondrium proliferation and neocartilage formation. These results suggested that angiogenesis may be important for the induction and differentiation of MSCs/cartilage precursor cells in vivo, and that morphological changes, once occurring, are maintained.

Intraarticularly-Injected Mesenchymal Stem Cells Stimulate Anti-Inflammatory Molecules and Inhibit Pain Related Protein and Chondrolytic Enzymes in a Monoiodoacetate-Induced Rat Arthritis Model.

Persistent inflammation is well known to promote the progression of arthropathy. mesenchymal stem cells (MSCs) have been shown to possess anti-inflammatory properties and tissue differentiation potency. Although the experience so far with the intraarticular administration of mesenchymal stem cell (MSC) to induce cartilage regeneration has been disappointing, MSC implantation is now being attempted using various surgical techniques. Meanwhile, prevention of osteoarthritis (OA) progression and pain control remain important components of the treatment of early-stage OA. We prepared a shoulder arthritis model by injecting monoiodoacetate (MIA) into a rat shoulder, and then investigated the intraarticular administration of MSC from the aspects of the cartilage protective effect associated with their anti-inflammatory property and inhibitory effect on central sensitization of pain. When MIA was administered in this rat shoulder arthritis model, anti-Calcitonin Gene Related Peptide (CGRP) was expressed in the joint and C5 spinal dorsal horn. Moreover, expression of A disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5), a marker of joint cartilage injury, was similarly elevated following MIA administration. When MSC were injected intraarticularly after MIA, the expression of CGRP in the spinal dorsal horn was significantly deceased, indicating suppression of the central sensitization of pain. The expression of ADAMTS 5 in joint cartilage was also significantly inhibited by MSC administration. In contrast, a significant increase in the expression of TNF-α stimulated gene/protein 6 (TSG-6), an anti-inflammatory and cartilage protective factor shown to be produced and secreted by MSC intraarticularly, was found to extend to the cartilage tissue following MSC administration. In this way, the intraarticular injection of MSC inhibited the central sensitization of pain and increased the expression of the anti-inflammatory and cartilage protective factor TSG-6. As the least invasive conservative strategies possible are desirable in the actual clinical setting, the intraarticular administration of MSC, which appears to be effective for the treatment of pain and cartilage protection in early-stage arthritis, may achieve these aims.

Involvement of necroptosis, a newly recognized cell death type, in steroid-induced osteonecrosis in a rabbit model.

We investigated the role of programmed necrosis (necroptosis), a newly recognized form of cell necrosis that has been implicated in the development of steroid-induced osteonecrosis. We used an osteonecrosis model in which 30 Japanese white rabbits each weighing 3.5kg were injected once with methylprednisolone at 20 mg/kg body weight into the right gluteal muscle. Ten animals killed 14 days thereafter were designated as S14d groups, while another 10 animals injected with necroptosis, a specific inhibitor of necrostatin-1 i.v. at 1.65mg/kg on the same day as the steroid were also killed on the 14th day and designated as SN14d group. As a control, 10 animals injected only with physiological saline were studied as N group. After the animals were sacrificed the bilateral femoral bone was examined histopathologically and the presence of osteonecrosis determined. Furthermore, animals subjected to the same treatment and killed on the 3rd day after drug administration were set up as S3d group and SN3d group, and Western blotting of Receptor-interacting protein ( RIP ) 1 and RIP3 in femoral bone performed. The osteonecrosis rate was 70% in S14d group, and 0% in both N and SN groups. In 2 of 10 animals in SN group fatty marrow was found. On Western blotting significantly increased expression of both RIP1 and RIP3 was noted in S3d group, confirming that Nec-1 was suppressed. Necroptosis mediated by RIP1 and RIP3 expression was thought to be implicated in the development of steroid-induced osteonecrosis. Also, by suppressing expression of RIP1 and 3 with the administration of Nec-1 the osteonecrosis rate was significantly decreased. These results suggest that necroptosis may have potential as a novel target for both elucidating the mechanisms underlying steroid-induced osteonecrosis and establishing more effective prophylactic countermeasures.

Prevention of glucocorticoid-associated osteonecrosis by intravenous administration of mesenchymal stem cells in a rabbit model.

BACKGROUND: Glucocorticoid-associated osteonecrosis is an intractable condition, making the establishment of preventative strategies of particular importance. Recently various studies using mesenchymal stem cells (MSC) have been conducted. Using a rabbit glucocorticoid-associated osteonecrosis model we administered green fluorescent protein (GFP)-labeled MSC intravenously to investigate their effect on osteonecrosis. METHODS: A rabbit osteonecrosis model in which methylprednisolone (MP) 20 mg/kg was injected into the gluteus of a Japanese white rabbit was used. Simultaneously with MP, MSC labeled with GFP (GFP-labeled MSC) were injected intravenously. Fourteen days later the animals were killed (MSC(+)/MP(+)/14d), femurs were extracted, and the prevalence of osteonecrosis was determined histopathologically. Also, animals were killed 3 days after simultaneous administration of GFP-labeled MSC and MP (MSC(+)/MP(+)/3d), and western blotting (WB) for GFP was performed of the femur, liver, kidney, lung, blood vessel, and vertebra, in addition to immunohistochemical study of femur. As a control for the histopathological study, animals were killed 14 days after MP administration and intravenous vehicle injection (MSC(-)/MP(+)/14d). For WB, animals were killed 3 days after intravenous GFP-labeled MSC administration and vehicle injection into the gluteus (MSC(+)/MP(-)/3d). RESULTS: In MSC(-)/MP(+)/14d osteonecrosis was found in 7 of 10 rabbits (70%), while in MSC(+)/MP(+)/14d, partial bone marrow necrosis was found in only 1 rabbit (12.5%); osteonecrosis was not found in 7 of 8 rabbits (p < 0.05). WB showed expression of GFP in the femur, not in the liver, kidney, lung, blood vessel, or vertebra, of MSC(+)/MP(+)/3d; expression of GFP-labeled MSC was absent in the femur of MSC(+)/MP(-)/3d. In the immunohistochemical study of MSC(+)/MP(+)/3d, homing of GFP-labeled MSC was noted perivascularly in the femur, but not in MSC(+)/MP(-)/3d. CONCLUSIONS: With transvenous MSC administration a significant prophylactic effect against glucocorticoid-associated osteonecrosis was found. Direct administration of MSC to the site of tissue injury requires highly invasive surgery. In contrast, as shown here the simple and hardly invasive intravenous administration of MSC may succeed in preventing osteonecrosis.

Osteocytic cell necrosis is caused by a combination of glucocorticoid-induced Dickkopf-1 and hypoxia.

Osteonecrosis is a major glucocorticoid-induced complication in the orthopedics field. Despite the extensive researches, mechanisms underlining the glucocorticoid-induced osteonecrosis are largely unknown. Here, we first provide the evidence that a combined treatment of cultured osteocytic cells with glucocorticoid and hypoxia caused necrotic cell death, which is assumed to occur in the acute bone injuries induced by glucocorticoids. We cultured MLO-Y4 murine osteocytic cells under hypoxia in the presence or absence of Dexamethasone (Dex) and examined the rates of apoptotic and necrotic cell death. Dex or hypoxia alone increased apoptotic cells, but not necrotic cells. The combination of Dex and hypoxia dramatically increased osteocytic cell death, notably necrotic cell death. The expression of Dickkopf-1 (Dkk-1), an inhibitor of Wnt/ß-catenin signal, was scarcely expressed in the control and hypoxic cells, but a dramatic increase of the Dkk-1 expression was detected in Dex-treated cells. siRNA-mediated knockdown of Dkk-1 in Dex and hypoxia-treated osteocytic cells showed the significant decreases in both apoptotic and necrotic cells. The results indicated that the combination of Dkk-1 overexpression by Dex and hypoxia causes the necrotic osteocytic cell death. The results also indicated that blocking of Dkk-1 can protect bone cells from glucocorticoid and hypoxia-induced cell injury.

Cryoglobulinemia vasculitis associated with adult-onset Still's disease.

Key Clinical Message: The present case indicates that cryoglobulinemia vasculitis should be considered in the differential diagnosis of purpura in patients with adult-onset Still's disease (AOSD). Abstract: The presence of purpura is suggested in adult-onset Still's disease (AOSD) hematological complications of hemophagocytic syndrome, disseminated intravascular coagulation, or thrombotic microangiopathy. We herein report a case of AOSD complicated by cryoglobulinemia vasculitis presenting with purpura.

Advantages of diffusion-weighted imaging over positron emission tomography-computed tomography in assessment of hilar and mediastinal lymph node in lung cancer.

BACKGROUND: The significance of diffusion-weighted imaging (DWI) is uncertain for the diagnosis of nodal involvement. The purpose of this study was to examine diagnostic capability of DWI compared with PET-CT for nodal involvement of lung cancer. METHODS: A total of 160 lung cancers (114 adenocarcinomas, 36 squamous cell carcinomas, and 10 other cell types) were analyzed in this study. DWI and PET-CT were performed preoperatively. RESULTS: The optimal cutoff values to diagnose metastatic lymph nodes were 1.70 × 10(-3) mm(2)/s for ADC value and 4.45 for SUVmax. DWI correctly diagnosed N staging in 144 carcinomas (90 %) but incorrectly diagnosed N staging in 16 (10 %) [3 (1.9 %) had overstaging, 13 (8.1 %) had understaging]. PET-CT correctly diagnosed N staging in 133 carcinomas (83.1 %) but incorrectly diagnosed N staging in 27 (16.8 %) [4 (2.5 %) had overstaging, 23 (14.4 %) had understaging]. Sensitivity, accuracy, and negative predictive value for N staging by DWI were significantly higher than those by PET-CT. Of the 705 lymph node stations examined, 61 had metastases, and 644 did not. The maximum diameter of metastatic lesions in lymph nodes were 3.0 ± 0.9 mm in 21 lymph node stations not detected by either DWI or PET-CT: 7.2 ± 4.1 mm in 39 detected by DWI, and 11.9 ± 4.1 mm in 24 detected by PET-CT. There were significant differences among them. The sensitivity (63.9 %) for metastatic lymph node stations by DWI was significantly higher than that (39.3 %) by PET-CT. The accuracy (96.2 %) for all lymph node stations by DWI was significantly higher than that (94.3 %) by PET-CT. CONCLUSIONS: DWI has advantages over PET-CT in diagnosing malignant from benign lymph nodes of lung cancers.

Relationship between vascular reactivity and expression of HMGB1 in a rat model of septic aorta.

INTRUODUCTION: High mobility group box 1 (HMGB1), a ubiquitous nuclear protein, induces several inflammatory diseases and functions as a fatal factor when released extracellularly. The effect of HMGB1 on vascular reactivity during sepsis remains to be clarified. METHODS: A rat model of abdominal sepsis was produced by cecal ligation and puncture (CLP) under sevoflurane anesthesia (n = 28). Anti-HMGB1 antibody at a dose of 4 or 0.4 mg/kg, or normal saline was injected twice intravenously, i.e., immediately after the CLP surgery and 4 h thereafter. Rats in the sham group underwent laparotomy, and the cecum was manipulated but not ligated or punctured. The descending thoracic aorta was excised 12 h after the CLP surgery and cut into rings of approximately 3 mm in length. Changes in the expression of HMGB1 and vascular reactivity were examined in the rings shortly after harvest and 4 h thereafter. RESULTS: HMGB1 was identified immunohistochemically and by Western blotting in the nuclei of vascular endothelial and smooth muscle cells in all groups shortly after excision of the aorta, but its expression was augmented only in the CLP groups 4 h thereafter. Degenerated smooth muscle cells were also observed after CLP. Anti-HMGB1 antibody dose-dependently inhibited the augmentation of HMGB1 expression and the morphological changes induced by CLP. The expression of HMGB1 partly correlated with suppression of vascular reactivity. CONCLUSION: The present results strongly suggest that HMGB1 plays an important role in vascular malfunction from an early phase of sepsis.

Solitary juvenile polyp of the rectum with intramucosal adenocarcinoma.

We describe a case of solitary juvenile polyp of the rectum with intramucosal adenocarcinoma. A 55-year-old man presented to our hospital for evaluation after a positive fecal occult blood test. Colonoscopy revealed a pedunculated polyp of 25 mm in size which has an irregular shape and pale red color on the rectum. The polyp had a proliferation of blood vessels and an invisible surface pattern. Endoscopic mucosal resection was performed. Pathologically, it was diagnosed as a solitary juvenile polyp with intramucosal well-differentiated adenocarcinoma. When we encounter juvenile polyps, the possibility of malignancy should be taken into consideration for treatment.

The Possibility of IPC to Prevent Ischemic-Reperfusion Injury in Skeletal Muscle in a Rat.

Blood removal with air tourniquets for a long time induces muscle damage after reperfusion. Ischemic preconditioning (IPC) has a protective effect against ischemia-reperfusion injury in striated muscle and myocardium. However, the mechanism of action of IPC on skeletal muscle injury is unclear. Thus, this study aimed to investigate the effect of IPC in reducing skeletal muscle damage caused by ischemia-reperfusion injury. The hindlimbs of 6-month-old rats were wounded with air tourniquets at a carminative blood pressure of 300 mmHg on the thighs. Rats were divided into the IPC (-) group and the IPC (+) group. The vascular endothelial growth factor (VEGF), 8-hydroxyguanosine (8-OHdG), and cyclooxygenase 2 (COX-2) were investigated by protein levels. Quantitative analysis of apoptosis was performed using the TUNEL method. Compared with the IPC (-) group, the IPC (+) group retained the VEGF expression, and the COX-2 and 8-OHdG expressions were suppressed. The proportion of apoptosis cells decreased in the IPC (+) group compared with the IPC (-) group. IPC in skeletal muscles proliferated VEGF and suppressed inflammatory response and oxidative DNA damage. IPC has the potential to reduce muscle damage after ischemia-reperfusion.