Mint3-mediated L1CAM expression in fibroblasts promotes cancer cell proliferation via integrin α5ß1 and tumour growth.

Fibroblasts are some of the major cells in tumour tissues that influence tumour progression and drug resistance. However, our understanding on fibroblast-mediated tumour malignancy remains incomplete. Munc18-1-interacting protein 3 (Mint3) is known as an activator of hypoxia-inducible factor-1 (HIF-1) even during normoxia in cancer cells, macrophages and fibroblasts. Although Mint3 promotes ATP production via glycolysis by activating HIF-1 in cancer cells and macrophages, the biological role of Mint3-mediated HIF-1 activation in fibroblasts remains unclear. To address this, we examined whether Mint3 in fibroblasts contributes to tumour growth. Mint3 depletion in mouse embryonic fibroblasts (MEFs) decreased tumour growth of co-injected human breast cancer cells, MDA-MB-231 and epidermoid carcinoma A431 cells in mice. In MEFs, Mint3 also promoted cancer cell proliferation in vitro in a cell-cell contact-dependent manner. Mint3-mediated cancer cell proliferation depended on HIF-1, and further gene expression analysis revealed that the cell adhesion molecule, L1 cell adhesion molecule (L1CAM), was induced by Mint3 and HIF-1 in fibroblasts. Mint3-mediated L1CAM expression in fibroblasts stimulated the ERK signalling pathway via integrin α5ß1 in cancer cells, and promoted cancer cell proliferation in vitro and tumour growth. In cancer-associated fibroblasts (CAFs), knockdown of MT1-MMP, which promotes Mint3-mediated HIF-1 activation, or Mint3 decreased L1CAM expression. As MEFs, CAFs also promoted cancer cell proliferation in vitro, and tumour growth via Mint3 and L1CAM. In human breast cancer specimens, the number of fibroblasts expressing L1CAM, Mint3 and MT1-MMP was higher in cancer regions than in adjacent benign regions. In addition, more phospho-ERK1/2-positive cancer cells existed in the peripheral region surrounded by the stroma than in the central region of solid breast cancer nest. Thus, Mint3 in fibroblasts might be a good target for cancer therapy by regulating cancer cell-stromal cell communication.

Utility of the FIB-4 Index for hepatocarcinogenesis in hepatitis C virus carriers with normal alanine aminotransferase levels.

The FIB-4 index is a simple formula using age, aspartate aminotransferase, alanine aminotransferase (ALT) and platelet count to evaluate liver fibrosis. We investigated the ability of the FIB-4 index for hepatocarcinogenesis in hepatitis C virus (HCV) carriers with normal ALT levels. A total of 516 patients with ALT levels persistently at or below 40 IU/L during an observation period of over 3 years were included. Factors associated with the development of HCC were determined. Hepatocellular carcinoma (HCC) developed in 60 of 516 patients (11.6%). The incidence rate of HCC at 5 and 10 years was 2.6% and 17.6%, respectively. When patients were categorized according to the FIB-4 index as ≤ 2.0 (n = 226), >2.0 and ≤ 4.0 (n = 169), and > 4.0 (n = 121), the cumulative incidence of HCC at 5 years was 0.5%, 1.3% and 8.0%, respectively, and 2.8%, 25.6% and 37.1% at 10 years, respectively. Patients with FIB-4 index >4.0 were at the highest risk (P < 0.001). Factors that were significantly associated with HCC in the multivariate analysis were FIB-4 index >2.0 (hazard ratio (HR), 7.690), FIB-4 index >4.0 (HR, 8.991), α-fetoprotein (AFP) >5 ng/mL (HR, 2.742), AFP >10 ng/mL (HR, 4.915) and total bilirubin >1.2 mg/dL (HR, 2.142). A scoring system for hepatocarcinogenesis that combines the FIB-4 index and AFP predicted patient outcomes with excellent discriminative ability. The FIB-4 index is strongly associated with the risk of HCC in HCV carriers with normal ALT levels.

Long-term efficacy and safety of sitagliptin in the treatment of Japanese Type 2 diabetes (ASSET-K1) to a target of HbA1c <7%.

BACKGROUND: Few studies have investigated the factors related to improvement and maintenance of glycemic control with sitagliptin in Type 2 diabetes (T2D) patients. AIM: To identify factors contributing to reaching and maintaining glycated hemoglobin (HbA1c) <7% with sitagliptin in Japanese T2D patients. SUBJECTS AND METHODS: This study included 1327 patients who were: taking sitagliptin as monotherapy; switched to sitagliptin; or taking sitagliptin in combination therapy. At baseline and 1, 3, 6, and 12 months after starting sitagliptin, weight, body mass index (BMI), HbA1c, fasting plasma glucose (FPG), and post-prandial plasma glucose (PPG) were measured. The subjects were divided into a group that achieved HbA1c<7% at 12 months, a poor control group (HbA1c≥8% at 12 months), and a discontinued group. Multiple regression analysis was performed to identify factors contributing to long-term control and maintenance with sitagliptin treatment. RESULTS: HbA1c decreased significantly from 8.0% at baseline to 7.3%, but weight was unchanged. FPG and PPG improved significantly. The HbA1c<7% group had a significantly higher age and a signifi cant ly lower BMI at baseline than the HbA1c≥8% group and the discontinued group. On multivariate regression analysis, baseline HbA1c, baseline BMI, and Δbody weight after 12 months were significantly related to HbA1c reduction. The most common adverse event was hypoglycemia, and the most common adverse event responsible for discontinuation was constipation. CONCLUSIONS: HbA1c<7.0% was achieved in 31% of T2D patients who had poor control with conventional treatment. Weight management is important for maintaining good long-term control with sitagliptin.

Correlation of serum ribavirin concentration with pretreatment renal function estimates in patients with chronic hepatitis C receiving combination antiviral therapy with peginterferon and ribavirin.

Serum ribavirin concentration is an important factor in antiviral therapy in combination with peginterferon (PEG-IFN) and ribavirin for patients with chronic hepatitis C in terms of both beneficial and adverse effects. We evaluated whether the serum ribavirin concentration can be predicted on the basis of renal function estimates. Serum creatinine and cystatin C concentrations were measured at the start of treatment in a total of 148 patients with chronic hepatitis C who underwent combination PEG-IFN and ribavirin therapy. Creatinine clearance (CrCl) and total clearance of ribavirin (CL/F) were calculated on the basis of the serum creatinine level. The glomerular filtration rate was calculated with two different formulae on the basis of the serum cystatin C level. These values were compared with serum ribavirin concentrations 4 weeks after the start of therapy. The cystatin C level increased with the progression of liver fibrosis, whereas the creatinine level was constant regardless of the degree of liver fibrosis. Significant correlation was not observed between the serum ribavirin concentration and serum creatinine level, cystatin C level, or calculated renal function estimates. However, significant correlation was found between the serum ribavirin concentration and CrCl and CL/F in patients who were given ribavirin >800 mg/day. Overall, renal function estimates do not correlate with the serum ribavirin concentration in Japanese patients with chronic hepatitis C who undergo combination PEG-IFN and ribavirin therapy. Serum creatinine-based renal function estimates might be predictive for the serum ribavirin concentration only in patients with a daily ribavirin intake of 800 mg or more.

Evaluation of optic nerve head configuration in various types of optic neuropathy with Heidelberg Retina Tomograph.

PURPOSE: To test whether Heidelberg Retina Tomograph (HRT) is applicable to assess the optic nerve head (ONH) configuration of the atrophic phase of non-glaucomatous optic neuropathy when a default set of the reference plane is used. METHODS: Ten eyes with non-arteritic anterior ischaemic optic neuropathy (NAION), 17 eyes with Leber's hereditary optic neuropathy (LHON), 40 eyes with compressive optic neuropathy (CON) owing to chiasmal tumour, and 241 eyes of control individuals were examined with HRT using the default reference plane. The global values of HRT parameters were evaluated among the groups of patients and controls. The sectoral measurements of the eyes with LHON and CON were compared with controls. To eliminate the influence of disc size and age on HRT measurements, eyes with disc area- and age-matched normal controls were used for comparison with eyes with NAION and LHON. RESULTS: Cup parameters in eyes with NAION were similar to those in controls. The retinal nerve fibre layer (RNFL) thickness was significantly thinner in eyes with NAION than that of controls. The eyes with LHON had significantly larger cup parameters, smaller rim volume, and thinner mean RNFL thickness than controls. Eyes with CON had significantly larger rim area and smaller cup parameters but similar RNFL thickness compared with controls. CONCLUSIONS: When the default reference plane is used, HRT can measure the ONH configuration in eyes with NAION and LHON as expected. However, caution must be made to interpret the parameters obtained from the eyes with CON.

Macular thickness reduction in eyes with unilateral optic atrophy detected with optical coherence tomography.

AIMS: To assess the changes in macular and peripapillary retinal nerve fibre layer (RNFL) thickness in eyes with unilateral optic atrophy and to evaluate the relationship between retinal thickness and visual function. METHODS: Enrolled were 22 patients with unilateral optic atrophy. Macular thickness at the divided nine areas and peripapillary RNFL thickness in quadrantic sections were measured by optical coherence tomography. Thickness values in the affected eyes were compared with those in the contralateral unaffected eyes. The correlation of foveal thickness with best-corrected visual acuity (BCVA) was evaluated. The correlation between retinal thickness and the remaining visual field area circumscribed with I-4-e isopter in superior and inferior hemifield was assessed. RESULTS: Macular thinning was observed in all areas (P < 0.001 in each area) other than the fovea (P = 0.068). Peripapillary RNFL thickness decreased in all quadrantic sections (P < 0.001 in each section). The affected to unaffected eye ratio of retinal thickness was more than 0.6 in each area. BCVA did not correlate with foveal thickness (correlation coefficient = 0.094, P = 0.668). Although not statistically significant (P = 0.281, superior hemifield; P = 0.053, inferior hemifield), there was a tendency that eyes with severe visual field loss show more marked retinal thinning. CONCLUSIONS: Macular thinning with the preserved foveal thickness is a hallmark of eyes with optic atrophy. Together with no correlation between foveal thickness and BCVA, this finding would help in differential diagnosis of macular and optic nerve diseases.

Germ cell-less expression in medaka (Oryzias latipes).

The gene germ cell-less (gcl) plays an important role in the early differentiation of germ cells in Drosophila. We isolated the gcl homolog of the model teleost medaka (Oryzias latipes) using degenerated primers and an ovary cDNA bank. The predicted amino acid sequence of medaka gcl showed 92, 68 and 31% overall identity to mouse, human and Drosophila gcl respectively. RT-PCR revealed stronger expression in the ovary and weaker expression in testis, brain, heart, liver and muscle tissue. Expression in early embryos indicates the presence of maternal mRNA. By in situ hybridisation (ISH), gcl could not be detected in embryos. In contrast to vasa, ISH revealed expression of gcl in the ovary but not in the testis. Mol. Reprod. Dev. 67: 15-18, 2004.

Gastrin and Helicobacter pylori in low-grade MALT lymphoma patients.

BACKGROUND: This study of patients with Helicobacter pylori infection and low-grade MALT lymphoma aimed to investigate: 1) the effect of H. pylori eradication therapy on the serum gastrin level, 2) whether changes of the serum gastrin level after therapy could predict the prognosis of patients with this tumour, and 3) the relationship between the gastric H. pylori load, the serum gastrin level and the status of MALT lymphoma. METHODS: Thirteen patients with documented low-grade MALT lymphoma and H. pylori infection were enrolled and received H. pylori eradication therapy as the sole initial treatment. The presence of H. pylori, the serum gastrin level, the endoscopic findings, the pathologic features of the biopsies and resected specimens, and the endoscopic ultrasonography findings were evaluated before and after therapy. Follow-up was carried out every 3-6 months. RESULTS: H. pylori eradication was eventually achieved in all 13 patients. The pretreatment fasting serum gastrin level decreased from 177.1 +/- 107.4 pg/ml to 129.2 +/- 78.1, 96.4 +/- 66.6 and 80.1 +/- 42.7 pg/ml after 0-3, 3-6 and 6-9 months, respectively (all P < 0.05). Successful eradication of H. pylori was followed by a decrease of the fasting serum gastrin level and complete regression of initial low-grade MALT lymphoma was observed in all patients. However, two patients subsequently developed recurrent high-grade MALT lymphoma or high-grade lymphoma. In one of them, the serum gastrin level rose again above the pretreatment value. In the other, however, the fasting gastrin level fell throughout the study period. The median fasting serum gastrin level before H. pylori eradication therapy was higher in the patients with tumours of the gastric body (203.4 +/- 108.9 pg/ml) than in those with tumours of the antrum and angulus (89.3 +/- 28.0 pg/ml) (P = 0.06). CONCLUSIONS: Hypergastrinaemia may be associated with an increased risk of gastric MALT lymphoma.

The position of the tibia during graft fixation affects knee kinematics and graft forces for anterior cruciate ligament reconstruction.

Ten cadaveric knees (donor ages, 36 to 66 years) were tested at full extension, 15 degrees, 30 degrees, and 90 degrees of flexion under a 134-N anterior tibial load. In each knee, the kinematics as well as in situ force in the graft were compared when the graft was fixed with the tibia in four different positions: full knee extension while the surgeon applied a posterior tibial load (Position 1), 30 degrees of flexion with the tibia at the neutral position of the intact knee (Position 2), 30 degrees of flexion with a 67-N posterior tibial load (Position 3), and 30 degrees of flexion with a 134-N posterior tibial load (Position 4). For Positions 1 and 2, the anterior tibial translation and the in situ forces were up to 60% greater and 36% smaller, respectively, than that of the intact knee. For Position 3, knee kinematics and in situ forces were closest to those observed in the intact knee. For Position 4, anterior tibial translation was significantly decreased by up to 2 mm and the in situ force increased up to 31 N. These results suggest that the position of the tibia during graft fixation is an important consideration for the biomechanical performance of an anterior cruciate ligament-reconstructed knee.

[MR imaging of posterior cruciate ligament injuries].

Posterior cruciate ligament(PCL) injuries are less frequent than anterior cruciate ligament(ACL) injuries, but are presumably more common than once thought. Thirty-nine patients with PCL injuries identified on MR images were studied. The criteria for PCL injury were complete tear, partial tear, and avulsion fracture. The approximate site of a partial tear was categorized as proximal, midsubstance, distal, or combination. Fourteen patients(35.9%) had complete tears of the PCL, 21 patients(53.8%) had partial tears, and four patients(10.3%) had avulsion fractures. A total of 12 patients (30.7%) had isolated PCL injuries, while the remaining 27 patients demonstrated evidence of other coexistent knee injuries, such as meniscal tears and ligamentous injuries. Of coexistent knee injuries, meniscal tears(18 patients, 46.2%) were most often seen.

[Three ocular sarcoidosis patients examined by indocyanine green angiography].

PURPOSE: To assess the findings of indocyanine green angiography(IA) in patients with ocular sarcoidosis. SUBJECTS AND METHODS: Three active ocular sarcoidosis patients with various retinochoroidal findings diagnosed by biopsy or systemic examination. Two patients were diagnosed pathologically and one patient was diagnosed clinically. IA & fluorescein angiography(FA) were performed before and after treatment with systemic steroid administration. RESULTS: IA revealed hyperfluorescence surrounding the presumed granulomatous lesions. This hyperfluorescence disappeared immediately after the treatment. FA showed hyperfluorescence continuing even after therapy. CONCLUSIONS: It is purposed that the ring-form hyperfluorescence in IA is due to accelerated vascular permeability in the active sarcoid granuloma. IA, which vividly reflects activity of sarcoid lesions, is an important tool for clinical evaluation of ocular sarcoidosis.

The biomechanical interdependence between the anterior cruciate ligament replacement graft and the medial meniscus.

To establish a quantitative biomechanical relationship between the anterior cruciate ligament graft and the medial meniscus, 10 human cadaveric knees were examined using the robotic/universal force-moment sensor testing system. In response to a combined 134-N anterior and 200-N axial compressive tibial load, the resulting kinematics of the knee and the in situ forces in the anterior cruciate ligament, the anterior cruciate ligament graft, and the medial meniscus were measured. Anterior tibial translation significantly increased after anterior cruciate ligament transection, between 6.8 +/- 2.3 mm at full extension and 12.6 +/- 3.3 mm at 30 degrees of flexion. Consequently, the resultant forces on the medial meniscus, ranging from 52 +/- 30 N to 63 +/- 51 N between full extension and 90 degrees of knee flexion in the intact knee, were doubled as a result of anterior cruciate ligament deficiency. However, after anterior cruciate ligament reconstruction, anterior tibial translations were restored to the levels of the intact knee, and thus the forces on the medial meniscus were restored as well. Likewise, the in situ forces in the anterior cruciate ligament replacement graft increased between 33% and 50% after medial meniscectomy.

P- and E-selectins recognize sialyl 6-sulfo Lewis X, the recently identified L-selectin ligand.

Recently we identified sialyl 6-sulfo Le(x) as a major L-selectin ligand on high endothelial venules of human peripheral lymph nodes. In this study we investigated the ligand activity of sialyl 6-sulfo Le(x) to E- and P-selectins and compared it with the binding activity of conventional sialyl Le(x), by using cultured human lymphoid cells expressing both carbohydrate determinants. The results of the recombinant selectin binding studies and the nonstatic monolayer cell adhesion assays indicated that both sialyl 6-sulfo Le(x) and conventional sialyl Le(x) served as ligand for E- and P-selectins, while L-selectin was quite specific to sialyl 6-sulfo Le(x). Anti-PSGL-1 antibodies as well as O-sialoglycoprotein endopeptidase treatment almost completely abrogated the binding of P-selectin but barely affected the binding of E-selectin, indicating that these carbohydrate determinants carried by O-glycans of PSGL-1 selectively serves as a ligand for P-selectin, while the ligand for E-selectin is not restricted to PSGL-1 nor to O-sialoglycoprotein endopeptidase-sensitive glycans. The binding of L-selectin was markedly reduced by O-sialoglycoprotein endopeptidase treatment but only minimally affected by anti-PSGL-1 antibodies, indicating that O-glycans carrying sialyl 6-sulfo Le(x) were the major L-selectin ligands, while PSGL-1 was only a minor core protein for L-selectin in these cells. These results indicated that each member of the selectin family has a distinct ligand binding specificity.

The forces in the anterior cruciate ligament and knee kinematics during a simulated pivot shift test: A human cadaveric study using robotic technology.

PURPOSE: Although it is well known that the anterior cruciate ligament (ACL) is a primary restraint of the knee under anterior tibial load, the role of the ACL in resisting internal tibial torque and the pivot shift test is controversial. The objective of this study was to determine the effect of these 2 external loading conditions on the kinematics of the intact and ACL-deficient knee and the in situ force in the ACL. TYPE OF STUDY: This study was a biomechanical study that used cadaveric knees with the intact knee of the specimen serving as a control. MATERIALS AND METHODS: Twelve human cadaveric knees were tested using a robotic/universal force-moment sensor testing system. This system applied (1) a 10-Newton meter (Nm) internal tibial torque and (2) a combined 10-Nm valgus and 10-Nm internal tibial torque (simulated pivot shift test) to the intact and the ACL-deficient knee. RESULTS: In the ACL-deficient knee, the isolated internal tibial torque significantly increased coupled anterior tibial translation over that of the intact knee by 94%, 48%, and 19% at full extension, 15 degrees, and 30 degrees of flexion, respectively (P <.05). In the case of the simulated pivot shift test, there were similar increases in anterior tibial translation, i.e., 103%, 61%, and 32%, respectively (P <.05). Furthermore, the anterior tibial translation under the simulated pivot shift test was significantly greater than under an isolated internal tibial torque (P <.05). Under the simulated pivot shift test, the in situ forces in the ACL were 83 +/- 16 N at full extension and 93 +/- 23 N at 15 degrees of knee flexion. These forces were also significantly higher when compared with those for an isolated internal tibial torque (P <.05). CONCLUSION: Our data indicate that the ACL plays an important role in restraining coupled anterior tibial translation in response to the simulated pivot shift test as well as under an isolated internal tibial torque, especially when the knee is near extension. These findings are also consistent with the clinical observation of anterior tibial subluxation during the pivot shift test with the knee near extension.

The effect of knee flexion angle and application of an anterior tibial load at the time of graft fixation on the biomechanics of a posterior cruciate ligament-reconstructed knee.

Ten knees were studied using a robotic testing system under a 134-N posterior tibial load at five flexion angles. Three knee positions were used to study the effect of flexion angle at the time of graft fixation (full extension, 60 degrees, and 90 degrees) and two were used to study the effect of anterior tibial load (60 degrees and 90 degrees). Knee kinematics and in situ forces were determined for the intact ligament and the graft for each reconstruction. Graft fixation at full extension significantly decreased posterior tibial translation compared with the intact knee by up to 2.9 +/- 2.9 mm at 30 degrees, while in situ forces in the graft were up to 18 +/- 35 N greater than for the intact ligament. Conversely, posterior tibial translation for graft fixation at 90 degrees was significantly greater than that of the intact knee by up to 2.2 +/- 1.1 mm at all flexion angles; in situ forces decreased as much as 33 +/- 30 N. When an anterior tibial load was applied before graft fixation at 90 degrees of flexion, posterior tibial translation did not differ from the intact knee from 30 degrees to 120 degrees, while the in situ force in the graft did not differ from the intact ligament at full extension, 60 degrees, and 120 degrees of flexion. These data suggest that graft fixation at full extension may overconstrain the knee and elevate in situ graft forces. Conversely, fixation with the knee in flexion and an anterior tibial load best restored intact knee biomechanics.

cDNA cloning of putative rat acetyl-CoA transporter and its expression pattern in brain.

Rat acetyl-CoA transporter gene (Acatn) encodes a hydrophobic multi-transmembrane protein involved in the O-acetylation of gangliosides. O-acetylated gangliosides have been found to play important roles in the embryonic development of the nervous system. We have isolated rat Acatn cDNA by PCR cloning. The amino acid sequence of rat Acatn exhibited 92% and 96% homology with human and mouse sequences, respectively. The mRNA was expressed in brain at all developmental stages. Acatn expression was higher in embryonic and postnatal rats than in adult rats. Cellular localization of Acatn mRNA in adult rat brain was also analyzed by in situ hybridization. Acatn mRNA expression was detected in the neuronal cells of cerebellum, hippocampus, hypothalamus, cortex, olfactory bulb, and dorsal and ventral anterior olfactory nucleus in adult rat brain.