Use of Cryopreserved Osteogenic Matrix Cell Sheets for Bone Reconstruction

Skeletal diseases, such as nonunion and osteonecrosis, are now treatable with tissue engineering techniques. Single cell sheets called osteogenic matrix cell sheets (OMCSs) grown from cultured bone marrow-derived mesenchymal stem cells show high osteogenic potential;however, long preparation times currently limit their clinical application. Here, we report a cryopreservation OMCS transplantation method that shortens OMCS preparation time. Cryopreserved rat OMCSs were prepared using slow- and rapid-freezing methods, thawed, and subsequently injected scaffold-free into subcutaneous sites. Rapid- and slow-frozen OMCSs were also transplanted directly to the femur bone at sites of injury. Slow-freezing resulted in higher cell viability than rapid freezing, yet all two cryopreservation methods yielded OMCSs that survived and formed bone tissue. In the rapid- and slow-freezing groups, cortical gaps were repaired and bone continuity was observed within 6 weeks of OMCS transplantation. Moreover, while no significant difference was found in osteocalcin expression between the three experimental groups, the biomechanical strength of femurs treated with slow-frozen OMCSs was significantly greater than those of non-transplant at 6 weeks post-injury. Collectively, these data suggest that slow-frozen OMCSs have superior osteogenic potential and are better suited to produce a mineralized matrix and repair sites of bone injury.

Utility of tricalcium phosphate and osteogenic matrix cell sheet constructs for bone defect reconstruction

AIM: To determine the effects of transplanting osteogenic matrix cell sheets and beta-tricalcium phosphate(TCP) constructs on bone formation in bone defects.METHODS: Osteogenic matrix cell sheets were prepared from bone marrow stromal cells(BMSCs), and a porous TCP ceramic was used as a scaffold. Three experimental groups were prepared, comprised of TCP scaffolds(1) seeded with BMSCs;(2) wrapped with osteogenic matrix cell sheets; or(3) both. Constructs were implanted into a femoral defect model in rats and bone growth was evaluated by radiography, histology, biochemistry, and mechanical testing after 8 wk. RESULTS: In bone defects, constructs implanted with cell sheets showed callus formation with segmentalor continuous bone formation at 8 wk, in contrast to TCP seeded with BMSCs, which resulted in bone nonunion. Wrapping TCP constructs with osteogenic matrix cell sheets increased their osteogenic potential and resulting bone formation, compared with conventional bone tissue engineering TCP scaffolds seeded with BMSCs. The compressive stiffness(mean ± SD) values were 225.0 ± 95.7, 30.0 ± 11.5, and 26.3 ± 10.6 MPa for BMSC/TCP/Sheet constructs with continuous bone formation, BMSC/TCP/Sheet constructs with segmental bone formation, and BMSC/TCP constructs, respectively. The compressive stiffness of BMSC/TCP/Sheet constructs with continuous bone formation was significantly higher than those with segmental bone formation and BMSC/TCP constructs.CONCLUSION: This technique is an improvement over current methods, such as TCP substitution, and is useful for hard tissue reconstruction and inducing earlier bone union in defects.

Indicators of sorafenib efficacy in patients with advanced hepatocellular carcinoma

AIM:To determine significant indicators for the efficacy of sorafenib in patients with advanced hepatocellular carcinoma(HCC).METHODS:A total of 46 patients with Barcelona Clinic Liver Cancer stage C who received sorafenib for more than 30 d at the Iizuka Hospital from June 2009 to December 2012 were enrolled in this study.Multivariate and univariate analyses were performed to evaluate the associations of hepatic function according to Child-Pugh grade,location and size of the largest tumor and adverse events of sorafenib treatment,such as hand-foot syndrome(HFS),hypertension,diarrhea,and alopecia,with the efficacy of treatment,as measured by overall survival(OS)and time to progression(TTP).RESULTS:Patients included 39 men and 7 women whose ages ranged from 48 to 85 years(70.6±9.6years).HCC was classified according to etiology as follows:hepatitis C virus(n=26),hepatitis B virus(n=9),and other(n=11).Liver function in patients was categorized as Child-Pugh grade A(n=30)or B(n=16).Tumors were categorized by size[<5 cm(n=33)or>5 cm(n=13)]and the location of the largest tumor was used to categorize patients with intrahepatic(n=28)or extrahepatic(n=18)HCC.HFS,hypertension,diarrhea,and alopecia were present in22(47.8%),19(41.3%),15(32.6%)and 7 patients(15.2%),respectively.The median OS of all patients was 373 d and the median TTP was 112 d.The etiology of HCC did not correlate with the median OS and TPP.The median OS of patients with tumors<5 cm was significantly longer than those with larger tumors(496 vs 245 d;HR=0.19,95%CI:0.07-0.48;P<0.01).According to the results of a multivariate analysis,the size of the largest tumor affected OS(HR=0.22,95%CI:0.08-0.59;P<0.01).The median TTP was significantly longer in patients with extrahepatic compared to intrahepatic major HCC(224 vs 98 d;HR=0.32;95%CI:0.14-0.67;P<0.01).The median TTP of patients with HFS was significantly longer than those without it(195 d vs 83 d;HR=0.41,95%CI:0.20-0.82;P<0.05),and the median TTP was significantly longer in patients with hypertension(195 d vs84 d;HR=0.43,95%CI:0.21-0.84;P<0.05).According to the results of the multivariate analysis,extrahepatic major HCC(HR=0.36,P<0.01)and HFS(HR=0.44,P<0.05)prolonged TTP.CONCLUSION:Extrahepatic major HCC and HFS are associated with prolonged TTP and are useful indicators for judging the efficacy of sorafenib treatment.

Multicentric recurrence of intraductal papillary neoplasm of bile duct after spontaneous detachment of primary tumor:A case report

BACKGROUND Intraductal papillary neoplasm of the bile duct(IPNB)rarely recurs in a multicentric manner.We encountered a patient with multiple recurrences of the gastric subtype of IPNB one year after spontaneous detachment of the primary tumor during peroral cholangioscopy(POCS).CASE SUMMARY A 68-year-old woman on maintenance hemodialysis because of lupus nephritis had several cardiovascular diseases and a pancreatic intraductal papillary mucinous neoplasm(IPMN).She was referred to our department for dilation of the common bile duct(CBD)and a tumor in the lumen,detected using ultrasonography.She had no complaints,and blood tests of hepatobiliary enzymes were normal.Magnetic resonance cholangiopancreatography(MRCP)showed a papillary tumor in the CBD with a filling defect detected using endoscopic retrograde cholangiography(ERC).Intraductal ultrasonography revealed a papillary tumor and stalk at the CBD.During POCS,the tumor spontaneously detached with its stalk into the CBD.Pathology showed low-intermediate nuclear atypia of the gastric subtype of IPNB.After 1 year,follow-up MRCP showed multiple tumors distributed from the left hepatic duct to the CBD.ERC and POCS showed multicentric tumors.She was alive without hepatobiliary symptoms at least two years after initial diagnosis of IPNB.CONCLUSION The patient experienced gastric subtype of IPNB without curative resection.Observation may be reasonable for patients with this subtype.