Evaluation of the Glasgow Prognostic Score in patients receiving chemoradiotherapy for stage III and IV esophageal cancer.

High Glasgow Prognostic scores (GPSs) have been associated with poor outcomes in various tumors, but the values of GPS and modified GPS (mGPS) in patients with advanced esophageal cancer receiving chemoradiotherapy (CRT) has not yet been reported. We have evaluated these with respect to predicting responsiveness to CRT and long-term survival. Between January 2002 and December 2011, tumor responses in 142 esophageal cancer patients (131 men and 11 women) with stage III (A, B and C) and IV receiving CRT were assessed. We assessed the value of the GPS as a predictor of a response to definitive CRT and also as a prognostic indicator in patients with esophageal cancer receiving CRT. We found that independent predictors of CRT responsiveness were Eastern Cooperative Oncology Group (ECOG) performance status, GPS and cTNM stage. Independent prognostic factors were ECOG performance status and GPS for progression-free survival and ECOG performance status, GPS and cTNM stage IV for disease-specific survival. GPS may be a novel predictor of CRT responsiveness and a prognostic indicator for progression-free and disease-specific survival in patients with advanced esophageal cancer. However, a multicenter study as same regime with large number of patients will be needed to confirm these outcomes.

BXSB-type genome causes murine autoimmune glomerulonephritis: pathological correlation between telomeric region of chromosome 1 and Yaa.

The autoimmune-prone BXSB/MpJ-Yaa mouse is a model of membranous proliferative glomerulonephritis (MPGN). Severe MPGN has been reported only in male BXSB/MpJ-Yaa mice because of the Y-linked autoimmune accelerator (Yaa) locus. However, we show that female BXSB/MpJ mice develop age-related MPGN without Yaa. Female BXSB/MpJ mice clearly developed MPGN characterized by increased mesangial cells, thickening of the glomerular basement membrane (GBM), double contouring and spike formation of GBM with T-cell infiltrations and podocyte injuries corresponding with increased autoantibody production and albuminuria. Analysis of the renal levels of the Fc gamma receptor (Fcgr) and interferon-activated gene 200 (Ifi200) family genes, which are MPGN candidate genes localized to the telomeric region of chromosome 1 (Chr.1), showed that Fcgr2b levels decreased, whereas Fcgr3 and Ifi202b levels increased in female BXSB/MpJ mice compared with healthy C57BL/6 mice. Furthermore, in isolated glomeruli, microarray analysis revealed that Fcgr3, Fcgr4 and Ifi202b expression was higher in male BXSB/MpJ-Yaa mice than in male BXSB/MpJ mice. These findings indicate that the BXSB/MpJ-type genome causes age-related MPGN with significant contribution from the telomeric region of Chr.1, and Yaa enhances the expression of genes localizing to this locus, thereby leading to severe MPGN in male mice.

An ocular infection model using suckling hamsters inoculated with equine herpesvirus 9 (EHV-9): kinetics of the virus and time-course pathogenesis of EHV-9-induced encephalitis via the eyes.

By using a new member of the neurotropic equine herpesviruses, EHV-9, which induced encephalitis in various species via various routes, an ocular infection model was developed in suckling hamsters. The suckling hamsters were inoculated with EHV-9 via the conjunctival route and were sacrificed after 6, 12, 24, 36, 48, 72, 96, 120, and 144 hours (h) post inoculation (PI). Three horizontal sections of the brains, including the eyes and cranial cavity, were examined histologically to assess the viral kinetics and time-course neuropathological alterations using a panoramic view. At 6 to 24 h PI, there were various degrees of necrosis in the conjunctival epithelial cells, as well as frequent mononuclear cell infiltrations in the lamina propria and the tarsus of the eyelid, and frequent myositis of the eyelid muscles. At 96 h PI, encephalitis was observed in the brainstem at the level of the pons and cerebellum. EHV-9 antigen immunoreactivity was detected in the macrophages circulating in the eyelid and around the fine nerve endings supplying the eyelid, the nerves of the extraocular muscles, and the lacrimal glands from 6 h to 144 h PI. At 96 h PI, the viral antigen immunoreactivity was detected in the brainstem at the level of the pons and cerebellum. These results suggest that EHV-9 invaded the brain via the trigeminal nerve in addition to the abducent, oculomotor, and facial nerves. This conjunctival EHV-9 suckling hamster model may be useful in assessing the neuronal spread of neuropathogenic viruses via the eyes to the brain.

Hapten-sandwich labeling. I. A general procedure for simultaneous labeling of multiple cell surface antigens for fluorescence and electron microscopy.

A hapten-sandwich procedure has been developed for specific labeling of cell surface antigens for fluorescence or electron microscopy. Haptens are azo-coupled to immunoglobulins specific for a cell surface antigen; the hapten-modified cell-bound antibodies can then be visualized by adding fluorescent antihapten antibody, or by adding antihapten antibody followed by hapten-modified markers for electron microscopy. Virus or high molecular weight protein markers are lightly cross-linked before conjugation with hapten to prevent their disruption. Such stable hapten-modified markers, and the accessibility of many different purified anti-azophenyl-hapten antibodies, make it feasible to distinguish more than one membrane antigen in a given labeling experiment. When mouse lymphoid cell populations are labeled with separate markers for Ig and for thymus-associated antigens, many cells exhibit the Ig marker exclusively or the thymic marker predominantly, and some cells are completely free of label.

Deuterium oxide: inhibition of calcium release in muscle.

Calcium release, measured as luminescence of the protein aequorin, was measured simultaneously with membrane potential and isometric tension in single muscle fibers of the barnacle (Balanus nubilus). Deuterium oxide inhibited calcium release and isometric tension but did not affect membrane potential, a result consistent with the postulate that deuterium oxide inhibits the coupling between excitation and contraction.

Involvement of organic anion transporting polypeptides in the toxicity of hydrophilic pravastatin and lipophilic fluvastatin in rat skeletal myofibres.

BACKGROUND AND PURPOSE: There is a discrepancy in the adverse effect of 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors, statins between the clinical reports and the studies using skeletal muscle cell models. In the clinical reports, both hydrophilic and lipophilic statins induce myotoxicity, whereas in in vitro experiments using cell lines of myoblasts, lipophilic, but not hydrophilic, statins exert myotoxicity. We investigated the cause of this discrepancy. EXPERIMENTAL APPROACH: Skeletal myofibres, fibroblasts and satellite cells were isolated from rat flexor digitorum brevis (FDB) muscles. Using these primary cultured cells as well as the L6 myoblast cell line, we compared the toxicity of hydrophilic pravastatin and lipophilic fluvastatin. The mRNA expression levels of possible drug transporters for statins were also examined in these cells using reverse transcriptase-PCR. KEY RESULTS: In the skeletal myofibres, both pravastatin and fluvastatin induced vacuolation and cell death, whereas in the mononuclear cells only fluvastatin, but not pravastatin, was toxic. mRNA of the organic anion transporting polypeptides (Oatp) 1a4 and Oatp2b1 were expressed in the skeletal myofibres, but not in mononucleate cells. Estrone-3-sulphate, a substrate for Oatps, attenuated the effects of pravastatin and fluvastatin in skeletal myofibres; p-aminohippuric acid, a substrate for the organic anion transporters (Oats), but not Oatps, failed to do so. CONCLUSIONS AND IMPLICATIONS: The statin transporters Oatp1a4 and Oatp2b1 are expressed in rat skeletal myofibres, but not in satellite cells, fibroblasts or in L6 myoblasts. This is probably why hydrophilic pravastatin affects skeletal muscle, but not skeletal myoblasts.

Effect of motion imagery to counter rest-induced suppression of F-wave as a measure of anterior horn cell excitability.

OBJECTIVE: To test if motor imagery prevents the rest-induced suppression of anterior horn cell excitability. METHODS: Ten healthy subjects underwent two separate experiments, each consisting of stimulating the median nerve 100 times and recording F-waves from abductor pollicis brevis (APB) in three consecutive sessions: (1) after muscle exercise to standardize the baseline, (2) after immobilization of APB for 3h and (3) after muscle exercise to check recovery. We instructed the subject to volitionally relax APB in experiment 1 (relaxation task), and to periodically simulate thumb abduction without actual movement in experiment 2 (imagery task). RESULTS: F-wave persistence and amplitude declined after relaxation task and recovered quickly after exercise, but changed little with imagery task. F-wave latencies showed no change when analyzed individually. The frequency distribution of collective F-waves recorded from all subjects remained the same after relaxation task, but showed a shift toward longer latencies after imagery task. CONCLUSIONS: Mental imagery without overt motor output suffices to counter the effect of sustained volitional muscle relaxation, which would, otherwise, cause a reversible reduction in anterior horn cell excitability. SIGNIFICANCE: This finding documents the importance of central drive for spinal excitability, which affects F-wave studies of a paretic muscle.

A light and electron microscopical study of the Tragulid (mouse deer) placenta.

The Tragulidae are the living relics of the basal ruminant stock. They have a diffuse placenta, with no aggregations of the placental villi into localised placentomes characteristic of all other ruminants. Despite this difference, this ultrastructural and immunocytochemical investigation demonstrates that in Tragulus the trophoblast binucleate cell (BNC) plays the same central role in development and structure as in all other ruminants. It shows an identical development and ultrastructure, produces granules reactive with bovine placental lactogen and pregnancy associated glycoprotein antibodies, and migrates when mature through the trophoblast tight junction to fuse into a mosaic of syncytial plaques from which the granules are released to the mother and which have replaced the uterine epithelium. Unlike the persistent plaques in the sheep and goat placenta, in Tragulus they are transient, dying by apoptosis with the fragments phagocytosed by the trophoblast. This brings the trophoblast into direct endotheliochorial apposition to maternal tissue until BNC migration and fusion replace the dead plaque. This intimate fetomaternal confrontation has not been shown in any other ruminant, and could be a relic of the evolutionary development of the synepitheliochorial from the original basic eutherian endo- or hemo-chorial placenta.

Delayed bile leakage from a remaining part of segment 8 in a posterior section graft after living donor liver transplantation: a common pitfall in harvesting a posterior section graft? A case report.

In Japan and Korea, where availability of deceased donor organs for solid organ transplantation remains rare, living donor liver transplantation (LDLT) using a posterior section graft (PSG; segments VI+VII, according to Couinaud's Nomenclature for liver segmentation) has now been accepted as a standard procedure that balances donor risk and patient benefits for cases in which right hemi-liver donation is too risky, because of marked volume imbalances between right and left hemi-livers. Compared with other types of grafts, however, the procedure requires detailed knowledge concerning hepatic vascular anatomy and meticulous manipulation during donation surgery. We present herein a case of delayed bile leakage from a remaining part of segment 8 in a PSG, which was considered to be a complication peculiar to LDLT using a PSG.

Increased risk of mammary carcinoma development following transplacental and trans-breast milk exposure to a food-derived carcinogen, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), in Sprague-Dawley rats.

Effects of transplacental and trans-breast milk exposure to a food-derived mammary and colon carcinogen, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), were investigated in rats. Female Sprague-Dawley rats were administered PhIP in the diet (100 ppm) for 4 weeks before mating with nontreated males and also during gestation and lactation. As controls, additional females were maintained on the basal diet without PhIP and mated as with the treated animals. The offspring of both groups were subdivided for each sex at weaning into three dietary groups receiving 100, 25, and 0 ppm and were killed at 47 weeks of age. Effects of the transplacental and neonatal exposure to PhIP on mammary carcinogenesis were most evident in females administered 25 ppm PhIP after weaning; the incidence and multiplicity of adenocarcinomas in offspring from the PhIP fed dams (42.9%, 0.62/rat) was significantly higher than the value for offspring from nontreated dams (4.8%, 0.05/rat). Furthermore, in the basal diet groups, the incidence of adenocarcinomas in females was higher, albeit not significantly, in offspring of the PhIP-treated than the nontreated dams (16.7%, 0.22/rat as compared with 3.3%, 0.07/rat). Although the highest incidence of mammary adenocarcinomas was found in the female progeny given 100 ppm PhIP from PhIP-treated dams (70.0%, 1.55/rat), this was only slightly higher than the 61.9% and 0.90/rat of the same dose group from the nontreated dams. In males, no apparent effects of transplacental and neonatal exposures were evident. In a separate experiment, excretion of PhIP into breast milk and transfer of PhIP to fetuses and neonates with resultant hepatic PhIP-DNA adduct formation were demonstrated. Thus, maternal exposure to this food-derived carcinogen may be a critical risk factor for generation of mammary carcinomas.

Frequent mutation of beta-catenin and APC genes in primary colorectal tumors from patients with hereditary nonpolyposis colorectal cancer.

Hereditary nonpolyposis colorectal cancer (HNPCC) is characterized by defective DNA mismatch repair, which results in genetic instability of tumors; however, only a few target genes have been recognized. Our previous study detected a low frequency of APC gene mutation (21%) in colorectal tumors from HNPCC patients, in contrast to a high frequency of APC gene alteration (>70%) in non-HNPCC tumors. Because both beta-catenin and ACP gene mutations have recently been shown to activate the same signaling pathway, we analyzed beta-catenin mutation in HNPCC tumors. A notable frequency of beta-catenin gene mutation (43%, 12 of 28) was found to occur in HNPCC colorectal tumors. Beta-catenin mutations were not detected in tumors with APC mutations. All beta-catenin mutations detected in HNPCC tumors existed within the regulatory domain of beta-catenin. Immunohistochemical staining of tumors with this mutation showed accumulation of beta-catenin protein in nuclei. These and previous data from our laboratory suggest that activation of the beta-catenin-Tcf signaling pathway, through either beta-catenin or APC mutation, contributes to HNPCC colorectal carcinogenesis in approximately 65% of cases.

Differential expression of small heat shock proteins in reactive astrocytes after focal ischemia: possible role of beta-adrenergic receptor.

Small heat shock proteins (sHSPs), a family of HSPs, are known to accumulate in the CNS, mainly in astrocytes, in several pathological conditions such as Alexander's disease, Alzheimer's disease, and Creutzfeldt-Jakob disease. sHSPs may act not only as molecular chaperones, protecting against various stress stimuli, but may also play a physiological role in regulating cell differentiation and proliferation. In the present study, we have demonstrated that transient focal ischemia in rats dramatically induced HSP27 but not alpha B-crystallin (alphaBC), both of which are members of sHSPs, in reactive astrocytes. In contrast, in vitro chemical ischemic stress induced both HSP27 and alphaBC in cultured glial cells to the same extent. Dibutyryl cAMP (dBcAMP) and isoproterenol, a beta-adrenergic receptor (betaAR) agonist, enhanced HSP27 expression but suppressed alphaBC, and changed the shape of the cells to a stellate form. dBcAMP and isoproterenol inhibited cell proliferation under normal conditions. An increase in betaAR-like immunoreactivity was also observed in reactive astrocytes in vivo. These results, together with recent findings that betaAR plays an important role in glial scar formation in vivo, raise the possibility that betaAR activation modulates sHSP expression after focal ischemia and is involved in the transformation of astrocytes to their reactive form.

Sodium-calcium exchange current. Dependence on internal Ca and Na and competitive binding of external Na and Ca.

Na-Ca exchange current was measured at various concentrations of internal Na [( Na]i) and Ca [( Ca]i) using intracellular perfusion technique and whole-cell voltage clamp in single cardiac ventricular cells of guinea pig. Internal Ca has an activating effect on Nai-Cao exchange beginning at approximately 10 nM and saturating at approximately 50 nM with a half maximum [Ca]i (Km[Ca]i) of 22 nM (Hill coefficient, 3.7). Measurement of Nai-Cao exchange current at various concentration of [Na]i revealed an apparent Km[Na]i of 20.7 +/- 6.9 mM (n = 14) with imax of 3.5 +/- 1.2 microA/microF. For [Ca]i transported by the exchange, a Km[Ca]i of 0.60 +/- 0.24 microM (n = 8) with an imax of 3.0 +/- 0.54 microA/microF was obtained by measuring Nao-Cai exchange current. These values are apparently different from the values for the external binding site which have been reported previously. Whether Na and Ca compete for the external binding site, and if so, how it affects the binding constants was then investigated. Outward Nai-Cao exchange current became larger by reducing [Na]o. The double reciprocal plot of the current magnitude and [Ca]o at different [Na]o revealed a competitive interaction between Na and Ca. In the absence of competitor [Na]o, an apparent Km[Ca]o of 0.14 mM was obtained. When comparing internal and external Km values, the external value is markedly larger than the internal one and thus we conclude that binding sites of the Na-Ca exchange molecule are at least apparently asymmetrical between the inside and outside of the membrane.

Translocation mechanism of Na-Ca exchange in single cardiac cells of guinea pig.

We have studied in single cardiac ventricular cells of guinea pig the ionic translocation mechanism of the electrogenic Na-Ca exchange, i.e., whether Na and Ca ions countercross the membrane simultaneously or consecutively with "ping pong" kinetics. The dose-response relation between the external Ca concentrations [( Ca]o) and the current density of the outward Na-Ca exchange current were measured at three different intracellular Na concentrations [( Na]i) in the absence of external Na. Nonlinear regression curves of the dose-response relation obtained by computer revealed Michaelis-Menten type hyperbola from which the [Ca]o giving a half-maximal response (apparent KmCao or K'mCao) and the apparent maximum current magnitude (I'max) were estimated at each [Na]i. As [Na]i increased, the K'mCao increased progressively and the value of K'mCao/I'max tended to decrease. These results are consistent with the simultaneous mechanism. The K'mCao/I'max values, however, were small and close to each other, so it was not possible to completely preclude a consecutive mechanism.

Comparative Studies on S-Glycoproteins Purified from Different S-Genotypes in Self-Incompatible BRASSICA Species II. Immunological Specificities.

Antisera were prepared by immunization of apparently purified S-glycoproteins; one from an S allele of Brassica campestris and two from S alleles of B. oleracea. Each antiserum was reactive not only with the homologous S-glycoprotein but also with the heterologous ones, i.e. with the S-glycoproteins of the other S alleles of the same locus. In double diffusion tests, a spur against the heterologous S-glycoproteins suggested heterogeneity of the glycoproteins. The heterogeneity appears to involve a component of the molecule in which the genotypic specificity of an S-glycoprotein resides, probably, for the recognition site. Some molecular components are common to all tested S-glycoproteins and in this respect are like the public antigens of the MHC locus of mammals. The common molecular components were recognized between the S-allele-specific glycoproteins within B. oleracea and also between them and those of B. campestris. No S-specific substances were detected in buffer soluble homogenates of style, ovary or anther. However, these homogenates contained substances that had structures similar to the corresponding common parts of the S-glycoproteins.

Circulating oxidized low density lipoprotein levels. A biochemical risk marker for coronary heart disease.

Recent studies have established oxidative modification of low density lipoprotein (LDL) as an important atherogenic factor. We examined the clinical relevance of circulating oxidized LDL (OxLDL) levels in atherosclerotic disease by an enzyme immunoassay with use of specific antibodies against OxLDL (FOH1a/DLH3) and apolipoprotein B. Plasma OxLDL levels were significantly higher in patients with coronary heart disease (n=65) than in control subjects (n=181; 201. 3+/-11.2 versus 112.4+/-3.3 U/dL, respectively; P<0.01). OxLDL levels were not associated with age, sex, total cholesterol, or apolipoprotein B levels in normal control subjects. Our results suggest that circulating OxLDL may be a possible biochemical risk marker for coronary heart disease.

Effects of FGFs on the morphogenic potency and AER-maintenance activity of cultured progress zone cells of chick limb bud.

The apical mesodermal region of chick limb buds (progress zone, PZ) which is essential for limb pattern formation contains uncommitted cells that change their positional values instructed by the apical ectodermal ridge (AER). Reciprically, the PZ cells maintain the AER activity. FGF-2 and FGF-4 can substitute for the AER to maintain normal outgrowth and gene expression in the limb bud. We examined the effects of FGF on the maintenance of PZ cells characteristics in culture by making recombinant limbs with anterior PZ cells that were pre-cultured in the presence of FGF-2 or FGF-4 and analyzed their morphogenic potency and responsiveness to positional cues arising from the zone of polarizing activity (ZPA). The limb buds expressed distal Hox genes and could form a segmented digit. Recombinant limb buds consisting of anterior PZ cells cultured without FGF failed to express Hox genes and formed instead a small cartilage nodule. These results indicate that addition of FGF-2 or FGF-4 to cultured PZ cells maintains their competence for Hox gene expression and digit formation, but not their responsiveness to positional cues from the ZPA. We also found that when anterior PZ cells which had been pre-cultured with FGF-2 or FGF-4 were implanted underneath the AER, they could maintain Fgf-8 expression in the AER, whereas this expression was not detected in the AER on the grafted anterior PZ cells that had been pre-cultured without FGF, indicating that FGF maintains AER-maintenance activity of PZ cells in culture.

Increased brain-derived neurotrophic factor-containing axons in the basal ganglia of patients with multiple system atrophy.

Brain-derived neurotrophic factor (BDNF) has a neurotrophic effect not only on mesencephalic dopaminergic neurons, but also on striatal neurons. To investigate whether the abnormal expression of BDNF occurs in the basal ganglia of patients with Parkinson disease (PD) and multiple system atrophy (MSA), we compared the BDNF levels in the striatum and globus pallidus of patients with PD or MSA to controls using immunohistochemistry. Furthermore, to quantitatively evaluate the immunohistochemical changes in the striatum, image analysis of the putamen was performed. BDNF-positive nerve fiber bundles and fine granular structures were scattered throughout the striatum and globus pallidus of all samples. Most of these granular structures were observed in glial fibrillary acidic protein-positive astrocytes. In addition, BDNF-positive neurites were abundant in the striatum of all MSA patients, and numerous BDNF-positive varicose fibers were found in the globus pallidus of some MSA cases with particularly severe striatal involvement. These observations suggest that the upregulated expression of BDNF may occur as a protective mechanism in the striatum of MSA patients, and that severe striatal degeneration may cause the aberrant accumulation of BDNF in the striatal projection areas of the globus pallidus of MSA patients.

Immunohistochemical localization of brain-derived neurotrophic factor in the spinal cords of amyotrophic lateral sclerosis and non-amyotrophic lateral sclerosis patients.

Brain-derived neurotrophic factor (BDNF) has a trophic effect on several neuronal subtypes including motor neurons. To localize and assess BDNF in the human spinal cord with particular reference to amyotrophic lateral sclerosis (ALS), we immunohistochemically studied spinal cords from 8 ALS and 13 non-ALS patients. Punctate staining for BDNF was observed in neuronal somata and proximal processes of large-sized anterior horn cells of non-ALS patients, as were distal axons immunolabeled in the neuropil. The same immunostaining pattern was found in the anterior horn cells of ALS patients. Neurons of the dorsal nucleus of Clarke, intermediolateral nucleus, and posterior horn sensory system were also stained in both groups. The results suggest that BDNF may act widely as a trophic factor in the human spinal cord, and motor neurons in ALS patients might be sufficiently supplied with endogenous BDNF from other neuronal subpopulations in the spinal cord.