Participation of hepatic α/ß-adrenoceptors and AT1 receptors in glucose release and portal hypertensive response induced by adrenaline or angiotensin II.

It has been previously demonstrated that the hemodynamic effect induced by angiotensin II (AII) in the liver was completely abolished by losartan while glucose release was partially affected by losartan. Angiotensin II type 1 (AT1) and adrenergic (∝1- and ß-) receptors (AR) belong to the G-proteins superfamily, which signaling promote glycogen breakdown and glucose release. Interactive relationship between AR and AT1-R was shown after blockade of these receptors with specific antagonists. The isolated perfused rat liver was used to study hemodynamic and metabolic responses induced by AII and adrenaline (Adr) in the presence of AT1 (losartan) and ∝1-AR and ß-AR antagonists (prazosin and propranolol). All antagonists diminished the hemodynamic response induced by Adr. Losartan abolished hemodynamic response induced by AII, and AR antagonists had no effect when used alone. When combined, the antagonists caused a decrease in the hemodynamic response. The metabolic response induced by Adr was mainly mediated by ∝1-AR. A significant decrease in the hemodynamic response induced by Adr caused by losartan confirmed the participation of AT1-R. The metabolic response induced by AII was impaired by propranolol, indicating the participation of ß-AR. When both ARs were blocked, the hemodynamic and metabolic responses were impaired in a cumulative effect. These results suggested that both ARs might be responsible for AII effects. This possible cross-talk between ß-AR and AT1-R signaling in the hepatocytes has yet to be investigated and should be considered in the design of specific drugs.

Participation of hepatic α/β-adrenoceptors and AT1 receptors in glucose release and portal hypertensive response induced by adrenaline or angiotensin II

It has been previously demonstrated that the hemodynamic effect induced by angiotensin II (AII) in the liver was completely abolished by losartan while glucose release was partially affected by losartan. Angiotensin II type 1 (AT1) and adrenergic (∝1- and β-) receptors (AR) belong to the G-proteins superfamily, which signaling promote glycogen breakdown and glucose release. Interactive relationship between AR and AT1-R was shown after blockade of these receptors with specific antagonists. The isolated perfused rat liver was used to study hemodynamic and metabolic responses induced by AII and adrenaline (Adr) in the presence of AT1 (losartan) and ∝1-AR and β-AR antagonists (prazosin and propranolol). All antagonists diminished the hemodynamic response induced by Adr. Losartan abolished hemodynamic response induced by AII, and AR antagonists had no effect when used alone. When combined, the antagonists caused a decrease in the hemodynamic response. The metabolic response induced by Adr was mainly mediated by ∝1-AR. A significant decrease in the hemodynamic response induced by Adr caused by losartan confirmed the participation of AT1-R. The metabolic response induced by AII was impaired by propranolol, indicating the participation of β-AR. When both ARs were blocked, the hemodynamic and metabolic responses were impaired in a cumulative effect. These results suggested that both ARs might be responsible for AII effects. This possible cross-talk between β-AR and AT1-R signaling in the hepatocytes has yet to be investigated and should be considered in the design of specific drugs.

Gait balance disorder by thalamic infarction with the disorder of interstitial nucleus of cajal.

The interstitial nucleus of Cajal (INC) is thought to play an important role in torsional/vertical eye position and head posture, and disorders of the INC induce abnormal ocular movements and head tilt. Our patients with ocular tilt reactions simultaneously also had disturbances in ambulatory balance, yet no reports address the loss of balance control induced by disorders of the INC. We examined the ambulatory disturbances induced by INC lesion. We experienced three patients with ocular movement disorders and abnormal head tilt due to thalamic infarction. We performed ophthalmic examinations on and checked the balance of them. With funduscopy, abnormal cycloduction was seen in the unaffected side and normal cycloduction was observed in the affected side. Nevertheless, Hess charts showed distortions in the visual image of both eyes. They all had disorders of balance control. We tried to treat them using the Bobath approach for improving their ambulatory balance. With subsequent improvements in balance control it was possible for them to take short walks, but it was difficult to make any improvements in their ocular movement. The INC is related to balance control of ambulation and disorders of the INC induce ambulatory disturbances. Cycloduction was only observed in the unaffected side, but Hess charts showed distortions of the visual image in both eyes. Ambulation was briefly improved, but diplopia persisted in these patients.

A component of polysaccharide peptidoglycan complex on Lactobacillus induced an improvement of murine model of inflammatory bowel disease and colitis-associated cancer.

Interleukin-6 (IL-6)/signal transducer and activator of transcription 3 (STAT3) signals play key roles in the pathogenesis of inflammatory bowel disease (IBD). We previously described that both intact cells and a cell wall-derived polysaccharide-peptidoglycan complex (PSPG) in a strain of lactobacillus [Lactobacillus casei Shirota (LcS)] inhibited IL-6 production in lipopolysaccharide (LPS)-stimulated lamina propria mononuclear cells (LPMCs) isolated from murine IBD. Diets with LcS improve murine IBD by suppression of IL-6 synthesis in LPMCs. Moreover, LcS supplementation with fermented milk ameliorates disease activity in patients with active ulcerative colitis. Here, we focused on the specific roles of PSPG in LcS concerning their anti-inflammatory actions. PSPG derived from LcS, and no other strain of lactobacilli, inhibited IL-6 production in LPS-stimulated murine IBD LPMCs. Purified PSPG-I from LcS inhibited IL-6 synthesis in LPS-stimulated murine IBD LPMCs through the inhibition of nuclear factor-kappaB. The anti-IL-6 action of LcS PSPG was abrogated by masking with monoclonal anti-PSPG-I. Furthermore, PSPG-I-negative L. casei strains (PSPG-I-negative mutant LcS: LC(DeltaPSPG-I), L. casei ATCC 334) did not inhibit IL-6 production. Finally, we confirmed the effects of PSPG-I on LcS in the models of both IBD and colitis-associated cancer (CAC). In the IBD model, ingestion of LcS improved ileitis and inhibited activation of IL-6/STAT3 signaling, while ingestion of the LC(DeltaPSPG-I) strain did not. In the CAC model, treatment with LcS, but not the LC(DeltaPSPG-I) strain, showed tumour-suppressive effects with an inhibition of IL-6 production in the colonic mucosa. These results suggested that a specific polysaccharide component in an L. casei strain plays a crucial role in its anti-inflammatory actions in chronic intestinal inflammatory disorders.

Triggering at the distal A2 pulley.

We report two cases with triggering at the distal end of the A2 pulley. One was caused by enlargement of the flexor digitorum profundus (FDP) tendon and the other by enlargement of both slips of the flexor digitorum superficialis (FDS) tendon. Both were relieved by reduction tenoplasty and short releases, or venting, of the distal A2 pulley.

Surface functionalization of inorganic nano-crystals with fibronectin and E-cadherin chimera synergistically accelerates trans-gene delivery into embryonic stem cells.

Stem cells holding great promises in regenerative medicine have the potential to be differentiated to a specific cell type through genetic manipulation. However, conventional ways of gene transfer to such progenitor cells suffer from a number of disadvantages particularly involving safety and efficacy issues. Here, we report on the development of a bio-functionalized inorganic nano-carrier of DNA by embedding fibronectin and E-cadherin chimera on the carrier, leading to its high affinity interactions with embryonic stem cell surface and accelerated trans-gene delivery for subsequent expression. While only apatite nano-particles were very inefficient in transfecting embryonic stem cells, fibronectin-anchored particles and to a more significant extent, fibronectin and E-cadherin-Fc-associated particles dramatically enhanced trans-gene delivery with a value notably higher than that of commercially available lipofection system. The involvement of both cell surface integrin and E-cadherin in mediating intracellular localization of the hybrid carrier was verified by blocking integrin binding site with excess free fibronectin and up-regulating both integrin and E-cadherin through PKC activation. Thus, the new establishment of a bio-functional hybrid gene-carrier would promote and facilitate development of stem cell-based therapy in regenerative medicine.

Endoscopic carpal tunnel release in the elderly.

The present study is aimed to clarify the postoperative outcome of endoscopic carpal tunnel release in elderly patients with carpal tunnel syndrome. Endoscopic carpal tunnel release was performed on 37 hands of 27 patients (2 men, 25 women) who were aged 70 years or older and clinically and electrophysiologically diagnosed with carpal tunnel syndrome. Mean age at the time of surgery was 74.5 years (range: 70-85 years). Mean postoperative follow-up was 35.5 months (range: 12-114 months). Pain was present preoperatively in 20 hands, but quickly resolved postoperatively in all cases. Numbness completely disappeared in 13 of 37 hands (35.1%), but some degree of numbness remained in the remaining cases. Preoperative severity of thenar muscle atrophy was none in 4 hands, mild in 7 hands, moderate in 12 hands and severe in 14 hands. Postoperative severity of thenar muscle atrophy at final follow-up was none in 13 hands, mild in 16 hands, moderate in 2 hands and severe in 6 hands, confirming that thenar muscle atrophy improves even in elderly patients. However, moderate or severe thenar muscle atrophy remained in 8 hands (21.6%). Endoscopic carpal tunnel release should be considered in the elderly, even though clinical symptoms may not improve substantially in advanced cases.

Induction of interleukin-12 by Lactobacillus strains having a rigid cell wall resistant to intracellular digestion.

Some strains of lactobacilli can stimulate macrophages and dendritic cells to secrete IL-12, which plays a key role in activating innate immunity. We examined the IL-12-inducing ability of 47 Lactobacillus strains belonging to 10 species in mouse peritoneal macrophages, and characterized the properties important for the induction of IL-12. Although considerable differences in IL-12-inducing ability were observed among the strains tested, almost all strains belonging to the Lactobacillus casei group (L. casei, Lactobacillus rhamnosus, and Lactobacillus zeae) or to Lactobacillus fermentum induced high levels of IL-12. Phagocytosis of lactobacilli was necessary for IL-12 induction, and the strains with strong IL-12 induction were relatively resistant to lysis in the macrophages. The sensitivity of Lactobacillus strains to in vitro treatment with M-1 enzyme, a member of the N-acetylmuramidases, was negatively correlated with IL-12-inducing ability. Using a probiotic strain, L. casei strain Shirota (LcS), we showed that the cell wall of LcS could be digested by long-term treatment with a high dose of M-1 enzyme and that the IL-12-inducing ability was diminished according to the duration of the enzyme treatment. The soluble polysaccharide-peptidoglycan complex released from the cell wall of LcS did not induce IL-12, whereas the insoluble intact cell wall of LcS induced IL-12. These results suggest that the intact cell wall structure of lactobacilli is an important element in the ability to induce IL-12 and that Lactobacillus strains having a rigid cell wall resistant to intracellular digestion effectively stimulate macrophages to induce IL-12.

pH-sensing nano-crystals of carbonate apatite: effects on intracellular delivery and release of DNA for efficient expression into mammalian cells.

Two unique and fascinating properties of carbonate apatite which are well-known in hard tissue engineering, have been unveiled, for the first time, for the development of the simplest, but most efficient non-viral gene delivery device - ability of preventing the growth of crystals needed for high frequency DNA transfer across a plasma membrane and a fast dissolution rate for effective release of DNA during endosomal acidification, leading to a remarkably high transgene expression (5 to 100-fold) in mammalian cells compared to the widely used transfecting agents. Moreover, by modulating the crystal dissolution rate of carbonate apatite through incorporation of fluoride or strontium into it, transfection activity could be dramatically controlled, thus shedding light on a new barrier in the non-viral route, which was overlooked so far. Thus we have developed an innovative technology with significant insights, that would come as a promising tool for both basic research laboratories and clinical settings.

Integrin-supported fast rate intracellular delivery of plasmid DNA by extracellular matrix protein embedded calcium phosphate complexes.

Extracellular matrix (ECM) proteins, such as collagen and fibronectin, play vital roles in development and maintenance of hard tissue (bone or tooth) and are, consequently, thoroughly investigated for construction of biomimetic scaffolds in combination with calcium phosphate (CaP) material (the major component of hard tissue) for bone or dental tissue engineering. Realizing the natural affinity of ECM components toward inorganic constituents of hard tissue, we successfully constructed the nanohybrids of DNA/CaP particles with either collagen 1 or fibronectin, which finally possessed the capability of specific recognition of integrin receptor for being swiftly internalized across the plasma membrane, leading to remarkably high transgene expression in mammalian cells. This new approach with precise receptor-specific delivery as well as 10- to 50-fold enhanced efficiency level compared to the classical one, has immediate applications for basic research and large scale production of recombinant therapeutic proteins and looks promising for gene therapy.

Role of E-cadherin molecules in spheroid formation of hepatocytes adhered on galactose-carrying polymer as an artificial asialoglycoprotein model.

The role of E-cadherin in the spheroid formation of hepatocytes adhered on the poly(N-p-vinylbenzyl-D-lactonamide) (PVLA) as a model ligand for asialoglycoprotein receptors (ASGP-R) of hepatocytes was studied. Expression of E-cadherin was increased in round hepatocytes adhered on a high-coating density of PVLA (100 microg/ml), and also in flat ones adhered on a low-coating density of PVLA (1 microg/ml) in the presence of epidermal growth factor (EGF). Hepatocyte spheroids formed on the high-coating density of PVLA in the presence of EGF after 48 h were inhibited by an anti-E-cadherin monoclonal antibody (ECCD-1). From immunofluorescence and confocal laser microscopy, E-cadherin was localized in the intercellular boundaries and concentrated at the inside surface of aggregated cells. As a result, E-cadherin could play an important role in hepatocyte assembly.

Probiotic Lactobacillus-induced improvement in murine chronic inflammatory bowel disease is associated with the down-regulation of pro-inflammatory cytokines in lamina propria mononuclear cells.

IL-6/STAT-3 signals play key roles in inflammatory bowel disease (IBD). It is known that Lactobacillus casei strain Shirota (LcS) improves inflammatory disorders. This study aimed to elucidate the effect of LcS on murine chronic IBD and to clarify the mechanism. We focused the inhibitory effect of LcS on the production of IL-6 in lipopolysaccharide (LPS)-stimulated large intestinal lamina propria mononuclear cells (LI-LPMC) isolated from mice with chronic colitis and in RAW264.7 cells in vitro. We also determined in vivo the effect of LcS on murine chronic IBD models induced with dextran sodium sulphate and SAMP1/Yit mice. Finally, we examined the cellular determinants of LcS for the down-regulation of IL-6 secretion by LI-LPMC, RAW264.7 cells and peripheral blood mononuclear cells (PBMC) derived from patients with ulcerative colitis (UC). LcS, but not other strains of Lactobacillus, inhibited the production of IL-6 in LPS-stimulated LI-LPMC and RAW264.7 cells, down-regulating the nuclear translocation of NF-kappaB. The LcS-diet-improved murine chronic colitis is associated with the reduction of IL-6 synthesis by LI-LPMC. LcS also improved chronic ileitis in SAMP1/Yit mice. The release of IL-6 in vitro in LPS-stimulated LI-LPMC, RAW 264.7 cells and UC-PBMC was inhibited by a polysaccharide-peptidoglycan complex (PSPG) derived from LcS. This probiotic-induced improvement in murine chronic inflammatory bowel disease is associated with the down-regulation of pro-inflammatory cytokines such as IL-6 and IFN-gamma production in LPMC. Therefore, LcS may be a useful probiotic for the treatment of human inflammatory bowel disease.

Fucoidan derived from Cladosiphon okamuranus Tokida ameliorates murine chronic colitis through the down-regulation of interleukin-6 production on colonic epithelial cells.

Our previous study indicated that the interleukin (IL)-6/STAT-3 signal was up-regulated in inflammatory bowel disease (IBD) in both humans and animal models. We also discovered phosphorylated STAT-3 in the nucleus of the colonic epithelial cells in IBD mice. Intestinal epithelial cells (IEC) have been shown to secrete IL-6. Therefore, the secretion of IL-6 from IEC may be one of the mechanisms of STAT-3 phosphorylation in IEC during the pathogenesis of IBD, and inhibition of IL-6 production by IEC may be beneficial in preventing IBD. We examined the preventative effect of various types of fucoidans on IL-6 production in a lipopolysaccharide (LPS)-stimulated murine colonic epithelial cells line, CMT-93, in vitro. We also determined in vivo the effect of fucoidans on murine chronic colitis induced with dextran sodium sulphate. Among fucoidans, those from Cladosiphon okamuranus Tokida and Kjellmaniella crassifolia inhibited IL-6 production in CMT-93 cells with the down-regulation of NF-kappaB nuclear translocation. Analysis of the effect of fucoidan on murine colitis in vivo showed that the disease activity index and myeloperoxidase activity decreased in mice fed Cladosiphon fucoidan, but not Fucus fucoidan. Cytokine profiles in colonic lamina propria indicated that the synthesis of interferon (IFN)-gamma and IL-6 decreased and that of IL-10 and transforming growth factor (TGF)-beta increased in mice fed Cladosiphon fucoidan, compared with mice fed a standard diet or Fucus fucoidan. The levels of IL-6 mRNA in colonic epithelial cells was lower in colitis-induced Balb/c mice fed Cladosiphon fucoidan than those fed a standard diet. Fucoidan improves murine chronic colitis by down-regulating the synthesis of IL-6 in the colonic epithelial cells. Fucoidan derived from C. o. Tokida may be useful as a dietary substance for the patients with inflammatory bowel disease.

[Multiple primary lung cancer diagnosed by thoracic computed tomography; report of 2 cases].

Two cases of bilateral multiple primary lung cancers are presented. One is synchronous, and another is metachronous. Case 1: An abnormal shadow in the left lower lobe was found on the chest radiograph of 40-year-old female during a regular check-up. Chest computed tomography (CT) film showed a micronodule in the right S2. Left lower lobectomy was performed, followed by second-staged right S2 segmentectomy 1 month after the initial operation. Case 2: A 62-year-old female underwent left lower lobectomy for lung cancer in April 1997. Follow-up CT showed a nodule in right S8. In March 2000, right basal segmentectomy was performed for second lung cancer. Moreover, follow-up CT showed a nodule in right S3. Right wedge resection was performed for third lung cancer in July 2002. It is important to carefully examine a synchronous lesion before the operation of a primary lung cancer and to perform close follow-up surveillance for early detection of a metachronous lesion.

Yeast pheromone signaling pathway as a bioassay to assess the effect of chemicals on mammalian peptide hormones.

The intercellular pheromone signal transduction pathways involved in sexual reproduction in the yeast Saccharomyces cerevisiae constitute an extracellular network system involving cell surface receptors. The system is analogous to the signaling pathway of mammalian peptide hormones. The yeast mating pheromone alpha factor is homologous to mammalian gonadoliberins such as luteinizing hormone-releasing hormone (LHRH). In this study, we used the yeast pheromone signaling pathway as a model system to evaluate the effect of industrial chemicals on mammalian peptide hormones. Haploid a- and alpha-cell types conjugate, using mating pheromones, to form diploid cells. However, in a cells treated with certain chemicals used in pesticides, fungicides, and industrial products (i.e., TPN (CAS No. 1897-45-6), thiuram (CAS No. 137-26-8), captan (CAS No. 133-06-2), oxine-copper (CAS No. 10380-28-6), zineb (CAS No. 12122-67-7), and ziram (CAS No. 137-30-4)) the induction of shmoo formation was suppressed even when commercial alpha-factor was added. The FUS1-lacZ gene, which is transcriptionally regulated by a pheromone, was transferred into yeast and the effects of TPN, captan, zineb, and ziram, under sublethal conditions, were investigated: beta-Galactosidase levels declined to levels similar to that of untreated control cells when in the absence of the alpha-factor. Furthermore, these chemicals influenced conjugation to alpha-cells, and mating efficiency declined as chemical concentration increased. Analysis of the yeast pheromone signaling pathway helps to establish chemical toxicity assay models for mammalian peptide signal transduction pathways.

Estimation of left ventricular systolic pressure by the left ventricular volume-time curve obtained from electrocardiograph gated 99m Tc-tetrofosmin single photon emission tomography using quantitative gated SPECT.

We report the estimation of left ventricular systolic pressure (LVSP) by a left ventricular (LV) volume-time curve obtained from electrocardiogram (ECG) gated 99mTc-tetrofosmin single photon emission computed tomography (SPECT) using quantitative gated SPECT (QGS). LVSP was calculated based on the following parameters: LV volumes, velocity and acceleration of LV contraction, aortic valve area and density of blood. The first three parameters can be derived from ECG gated SPECT. In 16 patients, the LV volume-time curve was obtained from ECG gated SPECT by using QGS. LVSP was estimated by the above-mentioned theory. The values of estimated peak LVSP were compared with those measured from a pressure transducer. There was a correlation between the values of peak LVSP estimated by the LV volume-time curve and those measured by pressure transducer (r=0.69, P<0.01). Using QGS, LVSP and the systolic LV pressure-volume relationship could be estimated by the LV volume-time curve.