Notch activation promotes osteoblast mineralization by inhibition of apoptosis.

Notch activator Jagged1 (JAG1) plays a critical role in the regulation of osteoblast differentiation and bone metabolism. In this study, JAG1-induced osteoblast proliferation, differentiation, and mineralization has been analyzed in primary osteoblasts for up to 7 days. Alkaline phosphatase and Alizarin red staining showed an enhanced osteoblast maturation and mineralization in JAG1 treated cells, as well as higher mRNA levels of late osteoblast differentiation markers. In contrast, Notch inhibitor DAPT and deletion of Runx2 totally blocked JAG1 effects on osteoblast mineralization. Flow cytometry data further showed a significantly higher cell proliferation in early stages of culture at day 3, and lower levels of osteoblast apoptosis in late stages of culture at day 7. More importantly, activation of anti-apoptotic factor BCL-2 was enhanced, while pro-apoptotic factor Caspase3 was reduced in JAG1 treated osteoblasts. Therefore, we conclude that cell mineralization is enhanced via anti-apoptotic actions of Notch signaling within the osteoblast cells.

Oleanolic Acid Enhances Mesenchymal Stromal Cell Osteogenic Potential by Inhibition of Notch Signaling.

Oleanolic acid (OA), a pentacyclic triterpenoid, has been shown to modulate multiple signaling pathways in a variety of cell linages. But the mechanisms underlying OA-mediated mesenchymal stromal cell (MSC) osteogenic differentiation are not known. In this study, we examined effects of OA on cell viability, osteogenic differentiation in MSCs, and the involvement of Notch and BMP signaling. OA induced bone marrow derived MSC differentiation towards osteoprogenitor cells and inhibited Notch signaling in a dose dependent manner. Constitutive activation of Notch signaling fully blocked OA induced MSC osteogenic differentiation. The expression level of early osteogenic marker genes, ALP, Runx2, and type I collagen, which play a critical role in MSC to osteoblast transition and servers as a downstream target of BMP signaling, was significantly induced by OA. Furthermore, BMP2 mediated MSC osteogenic differentiation was significantly enhance by OA treatment, indicating a synergistic effect between BMP2 and OA. Our results suggest that OA is a promising bioactive agent for bone tissue regeneration, and inhibition of Notch signaling is required for its osteogenic effects on MSCs.

Vacuum-assisted closure of necrotic and infected cranial wound with loss of dura mater: A technical note.

BACKGROUND: Complex cranial wounds can be a problematic occurrence for surgeons. Vacuum-assisted closure devices have a wide variety of applications and have recently been used in neurosurgical cases involving complex cranial wounds. There is only one report regarding the use of a vacuum-assisted closure device with loss of dura mater. We report a complicated case of a necrotic cranial wound with loss of dura mater. CASE DESCRIPTION: A 68-year-old female underwent an evacuation of a subdural hematoma. Postoperatively, the patient developed a wound infection that required removal of the bone flap. The wound developed a wedge-shaped necrosis of the scalp with exposure of brain tissue due to loss of dura mater from previous surgeries. She underwent debridement and excision of the necrotic tissue with placement of a synthetic dural graft (Durepair®, Medtronic, Inc.) and placement of a wound vac. The patient underwent a latissismus dorsi muscle flap reconstruction that subsequently failed. After the wound vac was replaced, the synthetic dural graft was replaced with a fascia lata graft and an anterolateral thigh free flap reconstruction. We describe the technical nuances of this complicated case, how the obstacles were handled, and the literature that discusses the utility. CONCLUSION: We describe a case of a complex cranial wound and technical nuances on how to utilize a wound-vac with loss of dura mater.

A prospective multivariate analysis of clinical factors associated with pouchitis after ileal pouch-anal anastomosis.

BACKGROUND & AIMS: Although acute pouchitis (AP) after ileal pouch-anal anastomosis (IPAA) for UC is common and easily treated, chronic pouchitis (CP) remains a difficult management issue. The aim of this study was to identify important clinical risk factors associated with AP or CP. METHODS: AP and CP were prospectively assessed, and demographic, disease, and treatment characteristics were tabulated. Univariate and multivariate analyses were performed to evaluate associations between AP or CP and potential risk factors. RESULTS: Two hundred IPAA patients were followed for a median of 24 months (range, 3-117 months). Thirty-six patients (18%) developed AP, and 23 patients (12%) developed CP. On univariate analysis, the use of steroids before colectomy and smoking were associated with the development of AP. CP was associated with male gender, smoking, length of follow-up, extraintestinal manifestations, backwash ileitis, and elevated (450x10(9)/L) platelet count. On multivariate analysis, the following risk factors were found to be independently associated with AP: use of steroids before colectomy (odds ratio [OR], 3.7; 95% confidence interval [CI], 1.5-8.9; P = .004) and smoking (OR, 2.3; 95% CI, 1.1-5.3; P = .04). CP was directly associated with extraintestinal manifestations (OR, 3.5; 95% CI, 1.1-11.1; P = .03), elevated platelet count (OR, 3.1; 95% CI, 1.1-8.9; P = .03), and increased length of follow-up (OR, 1.3; 95% CI, 1.1-1.6; P = .002). Smoking reduced the incidence of CP (OR, 0.2; 95% CI, 0.05-0.74; P = .04). CONCLUSIONS: Clinical factors associated with AP included use of steroids before colectomy and smoking. Factors directly related to CP were extraintestinal manifestations, elevated platelet count, and length of follow-up after IPAA. Smoking appears to protect against the development of CP.

The effect of antioxidants and a caspase inhibitor on cryopreserved rat hepatocytes.

Hepatocyte transplantation and use of bioartificial liver support systems have been suggested as potential therapies for fulminant hepatic failure. Cryopreservation in liquid nitrogen is presently the major method of long-term storage of isolated hepatocytes. However, cryopreservation can result in low cell recovery and reduction in differentiated function. Several possible mechanisms of cell death during cryopreservation have been proposed. The most important mechanisms appear to be oxidative stress and apoptosis. In this study, we isolated fresh rat hepatocytes and cryopreserved them in three media: University of Wisconsin (UW) solution, an antioxidant-containing medium, and medium containing a caspase inhibitor. Viability and function of hepatocytes cryopreserved in these media were examined. Cryopreservation conditions had no effect on hepatocyte viability after thawing. However, after culture we found significant improvements in viability and function in both antioxidant- and caspase inhibitor-treated hepatocytes at 6 and 24 h.

Transcriptomic fingerprinting of bone marrow-derived hepatic beta2m-/Thy-1+ stem cells.

The aim of the present study was to determine if the bone marrow (BM) beta2m-/Thy-1+ stem cells isolated from common bile duct ligated (CBDL) rats possess hepatocyte-like characteristics in their global gene expression profiles. The Affymetrix RG U34A arrays were used to conduct transcriptomic profiling on BM beta2m-/Thy-1+ stem cells isolated from CBDL and control rats as well as primary hepatocytes. Forty-one probe sets were up-regulated more than 2-fold in CBDL-derived beta2m-/Thy-1+ BM stem cells compared to control BM stem cells. Twenty-seven probe sets were present in both CBDL-derived beta2m-/Thy-1+ BM stem cells and control hepatocytes but absent in control beta2m-/Thy-1+ BM stem cells, including Tcf1 and Dbp. Compared to the control beta2m-/Thy-1+ BM stem cells, CBDL-derived beta2m-/Thy-1+ BM stem cells shared more commonly expressed genes with hepatocytes. Overall, CBDL-derived beta2m-/Thy-1+ stem cells displayed a different transcriptomic fingerprint compared with beta2m-/Thy-1+ BM stem cells isolated from control rats; and CBDL-derived beta2m-/Thy-1+ stem cells started to express some hepatocyte-like genes.

Central and peripheral actions of somatostatin on the growth hormone-IGF-I axis.

Somatostatin (SRIF) analogs provide safe and effective therapy for acromegaly. In a proportion of patients, however, SRIF analogs may lead to discordant growth hormone (GH) and IGF-I suppression, which suggests a more complex mechanism than attributable to inhibition of GH release alone. To elucidate whether SRIF acts peripherally on the GH-IGF-I axis, we showed that rat hepatocytes express somatostatin receptor subtypes-2 and -3 and that IGF-I mRNA and protein levels were suppressed in a dose-dependent manner by administration of octreotide. The inhibitory effect of SRIF was not apparent without added GH and in the presence of GH was specific for IGF-I induction and did not inhibit GH-induced c-myc or extracellular signal regulated kinase (ERK) phosphorylation. Pertussis toxin treatment of hepatocytes incubated with GH and SRIF, or with GH and octreotide, abrogated the inhibitory effect on GH-induced IGF-I, which confirms the requirement for the inhibitory G-protein. Treatment with SRIF and GH increased protein tyrosine phosphatase (PTP) activity and inhibited signal transducer and activator of transcription-5b (STAT5b) phosphorylation and nuclear localization. Octreotide also inhibited GH-stimulated IGF-I protein content of ex vivo-perfused rat livers. The results demonstrate that SRIF acts both centrally and peripherally to control the GH-IGF-I axis, providing a mechanistic explanation for SRIF analog action in treating patients with GH-secreting pituitary adenomas.

The value of laparoscopy in management of abdominal trauma.

The role of laparoscopy (LS) in abdominal trauma is controversial. Concerns remain regarding missed injuries and safety. Our objective for this study was to determine the safety and better define the role of LS in abdominal trauma victims. We performed a retrospective review of all patients who sustained abdominal trauma and underwent LS in a level I trauma center. The main outcome measures were age, gender, mechanism of injury (MOI), indication for laparoscopy, presence of intra-abdominal injury (IA), therapeutic laparoscopy (TxLS), need for laparotomy, length of hospital stay (LOS), missed injuries, complications, and deaths. Forty-eight patients underwent LS (62 per cent male; average age, 28 years; MOI, 35 (85%) penetrating, 7 (15%) blunt; mean ISS, 8). At laparoscopy, 58 per cent of patients had no intra-abdominal injury. IA injury was treated with laparotomy in 14 (29%) and TxLS in 6 (13%). One patient had a negative laparotomy (2%). No injuries were missed. No patients required reoperation. There was one complication: a pneumothorax. There were no deaths. LS was most valuable in penetrating trauma, avoiding laparotomy in more than two-thirds of patients with suspected intra-abdominal injury. LS can serve as a useful adjunct for the evaluation of blunt trauma. In a level I trauma center with LS readily available, the procedure is associated with a low rate of complications and missed injury.