Expression of a 1,25-dihydroxyvitamin D3 membrane-associated rapid-response steroid binding protein during human tooth and bone development and biomineralization.

The calciotropic hormone 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] has been established to control skeletal tissue formation and biomineralization via the regulation of gene expression. This action involves the well-characterized nuclear 1,25(OH)2D3 receptor. However, it has been recognized that several cellular responses to 1,25(OH)2D3 may not to be related to the exclusive nuclear receptor. Indeed, this secosteroid is able to generate rapid responses that have been proposed to be mediated by interactions of the ligand, which is a putative cell membrane-associated rapid-response steroid (MARRS) binding protein for 1,25(OH)2D3 [1,25D3-MARRS]. The nongenomic pathway of 1,25(OH)2D3 was studied here in detail by immunolocalization of the 1,25D3-MARRS during the specific context of human prenatal development. Western blotting with proteins extracted from 4 week- to 27-week-old embryos was performed, evidencing a 65-kDa molecular species recognized by antibody Ab 099 generated against synthetic peptides corresponding to the N terminus of the 1,25D3-MARRS from chick intestinal basolateral membranes. Based on this biochemical conservation of protein in the human species, the temporospatial expression patterns were established in the craniofacial skeleton at the same ages. Comparative analysis was performed in teeth and bones from early morphogenesis to terminal cell differentiation and extracellular biomineralization. The data show the potential implication of 1,25D3-MARRS in the heterogeneous cell population including ameloblasts, odontoblasts, osteoblasts, and osteoclasts. The epithelial-mesenchymal cascade related to odontogenesis was coincident with a sequence of up- and down-regulation of immunoreactive 1,25D3-MARRS. Biomineralization was associated with a striking up-regulation in the adjoining secretory cells in all tissues. Finally, osteoclasts appeared also to express the 1,25D3-MARRS during these early phases of bone modeling. Previously obtained data of the nuclear vitamin D receptor (VDR) expression and this study on 1,25D3-MARRS suggest the existence of cross-talk between the genomic and nongenomic pathways during human development.     

Adeno-associated virus vector-mediated systemic delivery of IFN-beta combined with low-dose cyclophosphamide affects tumor regression in murine neuroblastoma models.

PURPOSE: Type I IFNs (IFN-alpha/beta) have shown significant antitumor activity in preclinical models but limited efficacy and significant toxicity in clinical trials. We hypothesized that the antitumor activity of type I IFNs could be enhanced by chronic, low-dose systemic delivery and sought to test this in murine neuroblastoma models. EXPERIMENTAL DESIGN: Continuous liver-generated expression of human IFN-beta (hINF-beta) was achieved through a gene therapy-mediated approach using adeno-associated virus vectors encoding hIFN-beta (AAV hINF-beta). Orthotopic localized retroperitoneal and disseminated models of neuroblastoma were established using three different xenografts. Immunohistochemical analysis and ELISA were used to evaluate the antiangiogenic effect of therapy. RESULTS: The development of both localized orthotopic (retroperitoneal) and disseminated neuroblastoma was prevented in all mice expressing hINF-beta. Continued growth of established retroperitoneal tumors, treated with AAV hINF-beta as monotherapy, was significantly restricted, and survival for mice with established, disseminated disease was significantly prolonged following administration of AAV hINF-beta. Analysis of treated tumors revealed a significant antiangiogenic effect. Mean intratumoral vessel density was diminished and expression of the angiogenic factors vascular endothelial growth factor and basic fibroblast growth factor were both decreased. Finally, combination therapy in which AAV hIFN-beta was used together with low-dose cyclophosphamide resulted in regression of both established retroperitoneal and disseminated disease. CONCLUSIONS: AAV-mediated delivery of hIFN-beta when used as monotherapy was able to restrict neuroblastoma growth due in part to inhibition of angiogenesis. When used in combination with conventional chemotherapy, AAV hIFN-beta was able to effect complete tumor regression.     

CT图象的面罩式覆盖法定量诊断脂肪肝

目的　探讨脂肪肝的脂肪浸润程度的无创性检查方法及临床意义。方法　用 GE 30 0 0 I CT机扫描 36例脂肪肝患者肝脏、脾脏 ,用 CT机配备软件处理数据 ,面罩式覆盖法计算肝脏各个层面的脂肪面积与对应层面肝脏面积 ,并得出比值 (脂肪浸润指数 )。用生化仪检测肝功能指标。肝穿刺病理检测肝组织内脂肪浸润程度。结果　平均脂肪浸润指数与肝功能状态呈显著正相关 ,亦与肝穿刺的病理结果呈正相关 (r=0 .86 5 ,P<0 .0 1)。结论　 CT图象面罩式覆盖法对脂肪浸润的定量分析是诊断脂肪肝可靠的无创性检查方法 ,对非均匀性脂肪肝该方法优于肝穿刺     

Physiological analysis of motor reorganization following lower limb amputation.

It is now known that amputation results in reorganization of central motor pathways, but the mechanism for the changes is unclear. One possibility is alteration of the excitability of the alpha motoneurons. We studied motor reorganization and excitability of alpha motoneurons to Ia input in 6 subjects with unilateral lower limb amputation. A Cadwell MES-10 stimulator was used to deliver transcranial magnetic stimuli through a circular coil centered on the sagittal axis 4 cm anterior to Cz and through an 8-shaped coil positioned over scalp locations 1 cm apart along the coronal axis. Surface EMG was recorded bilaterally from quadriceps femoris, the first muscle immediately proximal to the site of amputation. Excitability of the spinal alpha motoneuron pool to Ia afferents was assessed by determining the ratio of the maximal H reflex to the maximal M response (H/M ratio) elicited in the quadriceps femoris. Stimuli of equal intensity delivered to optimal scalp positions recruited a larger percentage of the alpha motoneuron pool in muscles ipsilateral to the stump than in those contralateral to the stump (P less than 0.01). Mean onset latencies of motor evoked potentials were shorter in ipsilateral muscles than in contralateral muscles (P less than 0.01). Muscles ipsilateral to the stump showed a trend toward activation from a larger number of scalp positions than those contralateral to the stump (P = 0.06). There was no difference in the quadriceps H/M ratios (7.2% ipsilateral vs. 10.9% contralateral). The absence of changes in the excitability of the alpha motoneuron pool in the presence of motor reorganization targeting muscles proximal to the stump suggests that reorganization occurs proximal to the alpha motoneuron level.