Pathological analysis of batch safety testing of veterinary vaccines using small laboratory animals.

Batch safety tests (BSTs) of veterinary vaccines are conducted using small laboratory animals to assure the safety of vaccines according to several criteria, including clinical signs and change in body weight. Although the latter is used as an evaluation index in BSTs, there have been no reports on the internal changes that affect body weight during the test period. Therefore, we analyzed BST via pathological examination of the tested animals. Here, BSTs were performed for 176 batches using mice and 126 batches using of guinea pigs. Most of the gross findings could be classified into four lesion types (nodules, adhesions, ascites, no apparent signs), with only one vaccine inducing lesions that could not be classified into any of these four types. Histopathological examination revealed that the reactions caused by BST were pyogenic and/or granulomatous inflammation. Nodular or adhesive lesions comprised more severe pyogenic granulomatous inflammation than ascites or cases with no apparent gross lesions. These nodular or adhesive lesions were more frequently induced by vaccines that contained an adjuvant than by vaccines that did not contain an adjuvant. The cases with "exceptional" gross findings histologically presented severe necrosis of the hematopoietic system. Additional testing showed that these "exceptional" lesions were induced when a specific type of light liquid paraffin was injected along with other vaccine additives. Our results show that body weight loss and/or lesions during BST were induced by proinflammatory properties of the tested vaccines and that BST is a sensitive method for detecting unexpected effects of vaccine components.

Analysis of risk factors of postlaparoscopic shoulder pain.

AIM: Postlaparoscopic shoulder pain (SP), mainly caused by pneumoperitoneum with CO2 , sometimes suffers patients. This study was aimed to analyze the backgrounds of SP after gynecologic laparoscopy to clarify the risk of SP. METHODS: We analyzed answers of questionnaire about the degree of SP from 696 patients undergoing gynecologic laparoscopic surgery since 2014-2018. The questionnaire asks the degree of SP with numeric rating scale from 0 to 10, before and 3 days after operation. We defined cases in which postoperative score elevated more than three compared to preoperative score as SP(+). Analyzed backgrounds were age, parity, body mass index, operative method, operative duration and amount of hemorrhage. Statistics was performed by Fisher exact analysis as univariate analysis, and with logistic regression as multivariate analysis. All laparoscopic surgeries were performed under 10-12 mmHg in pressure of pneumoperitoneum with CO2 . RESULTS: Univariate analysis revealed categories 'less than 50 years old', and 'over 2 h' and 'over 3 h' in operative duration resulted significant high rate of SP(+). For these three factors, multivariate analysis resulted that "less than 50 years old' and 'over 3 h in operative duration' were significantly high. CONCLUSION: This study suggests that 'less than 50 years' old and 'over 3 h in operative duration' were risk factors of postlaparoscopic SP. To protect from SP after laparoscopy, some countermeasures should be necessary especially for these patients.

Complications Related to the Initial Trocar Insertion of 3 Different Techniques: A Systematic Review and Meta-analysis.

This systematic review aimed to investigate complications related to initial trocar insertion among 3 different laparoscopic techniques: Veress needle (VN) entry, direct trocar entry (DTE), and open entry (OE). A literature search was completed, and complications were assessed. Major vessel injury, gastrointestinal injury, and solid organ injury were defined as major complications. Minor complications were defined as subcutaneous emphysema, extraperitoneal insufflation, omental emphysema, trocar site bleeding, and trocar site infection. Arm-based network meta-analyses were performed to identify the differences in complications among the 3 techniques. Seventeen studies were included in the quantitative analysis. DTE resulted in fewer major complications when compared with VN entry although the difference was not significant (p = .23) as well as significantly fewer minor complications (p < .001). There were no significant differences in minor complications when comparing OE and DTE (p = .74). Fewer major complications were observed with OE compared with VN entry although the difference was not significant (p = .31). There were significantly fewer minor complications for patients who underwent OE (p = .01). DTE patients experienced the least number of minor complications followed by VN entry and OE. In conclusion, major complications are extremely rare, and all 3 insertion methods can be performed without mortality.

Second-generation total synthesis of aplyronine A featuring Ni/Cr-mediated coupling reactions.

Second-generation total synthesis of aplyronine A, a potent antitumor marine macrolide, was achieved using Ni/Cr-mediated coupling reactions as key steps. The overall yield of the second-generation synthetic pathway of aplyronine A was 1.4%, obtained in 38 steps based on the longest linear sequence. Compared to our first-generation synthetic pathway of aplyronine A, the second-generation synthesis greatly improved both the yield and number of steps. In particular, we improved the stereoselectivity in the construction of the C13 stereogenic center and the C14-C15 (E)-trisubstituted double bond using the asymmetric Ni/Cr-mediated coupling reaction. Furthermore, we established efficient reaction conditions for the asymmetric Ni/Cr-mediated coupling reaction between the C21-C28 segment and C29-C34 segment. Thus, this coupling reaction proceeded with an equimolar ratio of each segment.

Yucasin is a potent inhibitor of YUCCA, a key enzyme in auxin biosynthesis.

Indole-3-acetic acid (IAA), an auxin plant hormone, is biosynthesized from tryptophan. The indole-3-pyruvic acid (IPyA) pathway, involving the tryptophan aminotransferase TAA1 and YUCCA (YUC) enzymes, was recently found to be a major IAA biosynthetic pathway in Arabidopsis. TAA1 catalyzes the conversion of tryptophan to IPyA, and YUC produces IAA from IPyA. Using a chemical biology approach with maize coleoptiles, we identified 5-(4-chlorophenyl)-4H-1,2,4-triazole-3-thiol (yucasin) as a potent inhibitor of IAA biosynthesis in YUC-expressing coleoptile tips. Enzymatic analysis of recombinant AtYUC1-His suggested that yucasin strongly inhibited YUC1-His activity against the substrate IPyA in a competitive manner. Phenotypic analysis of Arabidopsis YUC1 over-expression lines (35S::YUC1) demonstrated that yucasin acts in IAA biosynthesis catalyzed by YUC. In addition, 35S::YUC1 seedlings showed resistance to yucasin in terms of root growth. A loss-of-function mutant of TAA1, sav3-2, was hypersensitive to yucasin in terms of root growth and hypocotyl elongation of etiolated seedlings. Yucasin combined with the TAA1 inhibitor l-kynurenine acted additively in Arabidopsis seedlings, producing a phenotype similar to yucasin-treated sav3-2 seedlings, indicating the importance of IAA biosynthesis via the IPyA pathway in root growth and leaf vascular development. The present study showed that yucasin is a potent inhibitor of YUC enzymes that offers an effective tool for analyzing the contribution of IAA biosynthesis via the IPyA pathway to plant development and physiological processes.

Azilsartan, an angiotensin II type 1 receptor blocker, restores endothelial function by reducing vascular inflammation and by increasing the phosphorylation ratio Ser(1177)/Thr(497) of endothelial nitric oxide synthase in diabetic mice.

BACKGROUND: Azilsartan, an angiotensin II type 1 (AT1) receptor blocker (ARB), has a higher affinity for and slower dissociation from AT1 receptors and shows stronger inverse agonism compared to other ARBs. Possible benefits of azilsartan in diabetic vascular dysfunction have not been established. METHODS: We measured vascular reactivity of aortic rings in male KKAy diabetic mice treated with vehicle, 0.005% azilsartan, or 0.005% candesartan cilexetil for 3 weeks. Expression of markers of inflammation and oxidative stress was measured using semiquantitative RT-PCR in the vascular wall, perivascular fat, and skeletal muscle. Phosphorylation of endothelial nitric oxide synthase (eNOS) at Ser1177 and Thr495 was measured using Western blotting, and the ratio of phosphorylation at Ser1177 to phosphorylation at Thr495 was used as a putative indicator of vascular eNOS activity. RESULTS: (1) Vascular endothelium-dependent relaxation with acetylcholine in KKAy mice was improved by azilsartan treatment compared to candesartan cilexetil; (2) the ratio of Ser1177/Thr495 phosphorylation of eNOS was impaired in KKAy and was effectively restored by azilsartan; (3) anomalies in the expression levels of monocyte chemotactic protein 1 (MCP1), F4/80, NAD(P)H oxidase (Nox) 2, and Nox4 of the aortic wall and in the expression of TNFα in the perivascular fat were strongly attenuated by azilsartan compared to candesartan cilexetil. CONCLUSIONS: These results provide evidence that azilsartan prevents endothelial dysfunction in diabetic mice, more potently than does candesartan cilexetil. Azilsartan's higher affinity for and slower dissociation from AT1 receptors may underlie its efficacy in diabetic vascular dysfunction via a dual effect on uncoupled eNOS and on Nox.

A small molecule inhibitor to plasminogen activator inhibitor 1 inhibits macrophage migration.

OBJECTIVE: Macrophage (Mϕ) migration rests on the adhesion/detachment between Mϕ surface components and extracellular matrixes, and the contribution of numerous inflammatory disorders. Plasminogen activator inhibitor (PAI)-1, a serine protease inhibitor, influences Mϕ motility through an action distinct from its classical modulation of the plasmin-based fibrinolytic process. We rely here on a small molecule PAI-1 inhibitor (TM5275) to investigate the role of PAI-1 in Mϕ migration in the pathogenesis of renal injury. APPROACH AND RESULTS: Mϕ migration was inhibited both in vitro and in vivo by TM5275. It was also reduced in T-cell-deficient nude mice, but not in PAI-1-deficient mice. Mϕ migration hinged on the interaction of PAI-1 with low-density lipoprotein receptor-related protein, an interaction prevented by TM5275, but not with vitronectin, urokinase-type plasminogen activator, or tissue-type plasminogen activator. Fed to rats with anti-Thy-1-induced nephritis, TM5275 significantly decreased Mϕ accumulation and ameliorated the progression of renal injury. CONCLUSIONS: These findings suggest that a small molecule PAI-1 inhibitor represents a novel class of anti-inflammatory agents targeting Mϕ migration by the inhibition of the interaction of PAI-1 with low-density lipoprotein receptor-related protein.

Effect of a dienogest for an experimental three-dimensional endometrial culture model for endometriosis.

The pathogenesis of endometriosis remains poorly understood at least in part because early stages of the disease process are difficult to investigate. Previous studies have proposed a three-dimensional fibrin matrix culture model to study human endometriosis. We examined the ultrastructural features of the endometriosis in this model and assessed the effect of a progestin on endometrial outgrowth and apoptosis in this culture system. Endometrial explants were placed in three-dimensional fibrin matrix culture and treated with and without various concentrations of the progestin dienogest. By the second week, endometrial gland-like formation was established in outgrowths both attached to and at a distance from the explants. These cells formed a combination of clumps and tubular monolayers surrounding a central cavity. Electron microscopy demonstrated that these cells are polarized with microvilli on the apical surface, desmosome-like structures, and basement membrane; features consistent with glandular epithelial cells. Outgrowth of endometrial stromal cells and glandular formation was impaired in response to dienogest in a dose-dependent manner. Our study shows that the human endometrial explants cultured in three-dimensional fibrin matrix establish outgrowths that ultrastructurally resemble ectopic endometrial implants. This model may provide insight into the cellular processes leading to endometriosis formation and enables screening of therapeutic compounds.

Effectiveness of upfront triple oral combination therapy with additional pirfenidone in a patient with severe pulmonary hypertension associated with lung diseases.

Diagnosis and treatment of pulmonary hypertension (PH) in patients with lung diseases (PH-LD) remain unestablished and pose significant challenges. In this report, we present a case of a 77-year-old patient with an indeterminate for usual interstitial pneumonia pattern along with chronic obstructive pulmonary disease, who developed groups 1 and 3 PH. Following diagnosis, upfront triple oral combination therapy (UTOCT) with macitentan, sildenafil, and selexipag was initiated. Stability in disease progression was achieved over 4 years with the addition of pirfenidone to address interstitial lung disease progression. To the best of our knowledge, this represents the first reported case of PH-LD, where disease control was maintained with the addition of pirfenidone to UTOCT. This case suggests that some patients with PH-LD, presenting with groups 1 and 3 PH, may benefit from combined UTOCT and antifibrotic agents, potentially improving symptoms and extending their prognosis.

Quality of End-of-Life in Cancer Patients With Dementia: Using A Nationwide Inpatient Database.

CONTEXT: The growing number of older people significantly affects end-of-life care. However, few studies have assessed the quality of end-of-life care among cancer patients with dementia. OBJECTIVES: To assess the quality of end-of-life care among non-small cell lung cancer patients with or without dementia using a nationwide inpatient database from Japan. METHODS: This was a retrospective observational study that used a nationwide inpatient database of 366 acute care hospitals from April 2014 to November 2018. Poisson regression models were used where the quality indicator was the dependent variable, dementia status was the independent variable, and the age group and Charlson comorbidity index were covariates. Incidence proportion ratios (IPRs) and confidence intervals (CIs) were obtained from the model. RESULTS: The study population included 16,758 patients, of whom 4507 (26.9%) had dementia. The incidence proportion of opioid use (61.8% vs. 70.8%; IPR: 0.87, 95% CI: 0.83-0.91), palliative care consultation (2.7% vs. 3.8%; IPR: 0.71, 95% CI: 0.58-0.88), mechanical ventilation (4.0% vs. 5.4%; IPR: 0.74, 95% CI: 0.62-0.87), and cardiopulmonary resuscitation (2.2% vs. 2.8%; IPR: 0.79, 95% CI: 0.63-0.99) was significantly lower in patients with dementia than in those without dementia. CONCLUSION: Patients with dementia are less likely to receive end-of-life care. This study demonstrates the importance of providing high-quality end-of-life care regardless the cognitive status of patients with cancer.

Bacterial production of the tunicate-derived antitumor cyclic depsipeptide didemnin B.

Natural products obtained from marine invertebrates such as sponges and tunicates are attractive sources of drugs. However, a critical obstacle in the development of these compounds is the problem of supply. In most cases, neither chemical synthesis nor mariculture of invertebrates is economically feasible. Due to structural similarities, many marine natural products are suspected to be produced by associated microorganisms. A favorable strategy for the production of such compounds is to use culturable microorganisms. Here we report that didemnin B, a tunicate-derived depsipeptide, has been isolated from a culturable bacterium, Tistrella mobilis YIT 12409.

Isolation and identification of cancer stem cells from a side population of a human hepatoblastoma cell line, HuH-6 clone-5.

PURPOSE: It has been thought that the persistence of even a small number of tumor cells in the body may increase each tumor cell in a similar manner and may allow the disease to proceed. However, only a few percent of such tumor cells exist in cancerous tissue. They are called "cancer stem cells (CSCs)". If an alternative method of annihilating CSCs is found, it will greatly deter relapse and metastasis. We attempted to identify and separate CSCs in hepatoblastoma aiming to develop a new therapy for hepatoblastoma. METHODS: The side population (SP) method was used as an indicator when extracting the CSC candidate group from the hepatoblastoma cells. The SP cells and non-SP cells were studied for tumourigenesis. RESULTS: Although tumors were formed when SP fraction cells were inoculated into mice, tumor formation was not observed in non-SP cells. SP cells had higher tumor formation ability compared to non-SP cells. CONCLUSION: Cancer stem-like cells were separated by the SP fraction method from hepatoblastoma cells. The in vivo experiment proved that SP fraction cells inoculated into mice were self-replicated, and the existence of cancer stem-like cells was identified.

Preliminary Effect and Acceptability of an Intervention to Improve End-of-Life Care in Long-Term-Care Facilities: A Feasibility Study.

The number of deaths of older adults in long-term care settings will increase with the aging population. Nurses and care workers in these settings face various challenges in providing end-of-life care, and interventions for quality end-of-life care may be useful. This feasibility study aims to explore the preliminary effect and acceptability of an intervention named the EOL Care Tool to improve end-of-life care in long-term-care facilities. We conducted a single-arm quasi-experimental study using mixed methods. This tool consisted of multiple components: professionalized lectures, newly developed structured documents, regular conferences regarding end-of-life care, and educational support from administrators. Twenty-four nurses and fifty-five care workers employed in a long-term care facility participated. For nurses, improvement in attitudes toward end-of-life care (p < 0.05) and interdisciplinary collaboration (p < 0.05) were shown quantitatively. Regarding acceptability, nurses and care workers evaluated the tool positively except for the difficulty of using the new documents. However, qualitative results showed that care workers felt the reluctance to address the work regarding end-of-life care. Therefore, a good preliminary effect and acceptability for nurses were indicated, while acceptability for care workers was only moderate. Revision to address the mentioned issues and evaluation of the revised tool with a more robust research design are required.

Pembrolizumab-induced secondary sclerosing cholangitis in a non-small cell lung cancer patient.

A 50-year-old woman with stage IV lung adenocarcinoma received seven cycles of pembrolizumab as third-line chemotherapy. Following the failure of pembrolizumab, she commenced fourth-line chemotherapy of docetaxel and ramucirumab. The patient complained of epigastric pain and a computed tomography (CT) scan revealed oedema-like thickening of the gallbladder wall, dilation of the bile ducts from the common to the intrahepatic bile ducts, and thickening of the common bile duct wall without any visible obstructions. Accumulation of fluorodeoxyglucose (FDG) in the gallbladder wall and bile duct was also detected with positron emission tomography (PET)-CT. A biopsy of the extrahepatic bile duct showed non-specific inflammation. Antibiotic treatment was not effective and pathogens were not detected. The patient was diagnosed with secondary sclerosing cholangitis (SSC) by pembrolizumab. She received 80 mg/day of prednisolone (PSL); however, SSC recurred with tapering of PSL. SSC then improved with steroid pulse therapy and subsequently 50 mg/day azathioprine and 80 mg/day PSL.

Laparoscopically Diagnosed and Treated Ruptured Metastatic Ovarian Tumor.

A 51-year-old woman visited our institution with a chief complaint of abdominal pain. Blood laboratory testing revealed a carcinoembryonic antigen level of 13.4 ng/mL. Magnetic resonance imaging revealed a massive pelvic mass with marked wall thickening, partly accompanied by a high-signal-intensity cystic component in T2-weighted images. The entire tumor had low-signal intensity in T1-weighted images. We diagnosed a ruptured ovarian tumor, and the patient underwent emergent laparoscopic left salpingo-oophorectomy. Pathological examination revealed metastatic colon cancer to the ovary, and lower gastrointestinal endoscopy confirmed sigmoid colon carcinoma. Laparoscopic sigmoidectomy was performed followed by adjuvant chemotherapy with capecitabine + oxaliplatin. Ruptured metastatic ovarian tumor is extremely rare. With early diagnosis and laparoscopic resection, the primary lesion can be identified and treated quickly.

Effects of rosuvastatin and colestimide on metabolic parameters and urinary monocyte chemoattractant protein-1 in type 2 diabetic patients with hyperlipidemia.

BACKGROUND: Rosuvastatin, a strong statin, and colestimide, a new anion exchange resin, are both clinically beneficial drugs for treatment of hypercholesterolemia. The main purpose of the study was to compare the effects of rosuvastatin and colestimide on metabolic parameters, adipokines, and markers of oxidative stress and diabetic nephropathy in patients with type 2 diabetes complicated by hyperlipidemia. DESIGN: A total of 40 patients with type 2 diabetes complicated by hyperlipidemia were recruited prospectively and consecutively. The patients were assigned randomly in equal numbers to rosuvastatin (2.5 mg/day) and colestimide (3.0 g/day) groups. Blood and urine tests were performed at the beginning of the study and after 12 weeks. RESULTS: Rosuvastatin significantly decreased the level of serum retinol-binding protein (RBP)-4, an insulin-resistant adipokine, in a subgroup of patients with poor glycemic control, in addition to exerting a strong low-density lipoprotein (LDL-C)-lowering effect. Colestimide significantly decreased HbA1c, even in patients treated with a sulfonylurea at a more than moderate dose, without influencing insulin resistance or adiponectin (an insulin-sensitive adipokine) and RBP4. Colestimide also significantly decreased the levels of urinary 8-iso-prostaglandin (PG) F2alpha (a marker of oxidative stress) and urinary monocyte chemoattractant protein-1 (MCP-1) (a marker of diabetic nephropathy). CONCLUSION: Our results show that rosuvastatin and colestimide exert different beneficial effects in type 2 diabetic patients complicated by hyperlipidemia. Therefore, concomitant use of these drugs may be useful for prevention of progression of diabetic complications.

Intraperitoneal administration of a small interfering RNA targeting nuclear factor-kappa B with paclitaxel successfully prolongs the survival of xenograft model mice with peritoneal metastasis of gastric cancer.

Activation of nuclear factor-kappa B (NF-kappaB) has been detected in various malignant tumors, including gastric carcinoma, and is associated with tumor growth, metastasis, resistance to chemotherapeutic agents and poor prognosis. Therefore, NF-kappaB is a potential target for antitumor therapy. In this study, we used a small interfering RNA (siRNA) to knockdown NF-kappaB p65 expression and determined whether intraperitoneal administration of NF-kappaB p65 siRNA and paclitaxel was effective for treating peritoneal metastasis of gastric cancer. Western blot analysis revealed that NF-kappaB p65 expression was diminished by NF-kappaB p65 siRNA. Apoptotic cells were increased after transfection of NF-kappaB p65 siRNA compared with control siRNA in the treatment with paclitaxel. In a murine xenograft model, abundant fluorescence was observed on the surface of intraperitoneal nodules of gastric cancer after siRNA administration. Moreover, intraperitoneal administration of NF-kappaB p65 siRNA reduced NF-kappaB expression in intraperitoneal nodules of gastric cancer. Finally, mice treated by intraperitoneal administration of NF-kappaB p65 siRNA and paclitaxel survived for a significantly longer time than mice treated by intraperitoneal administration of paclitaxel alone (p = 0.0002). Taken together, the present results demonstrate that intraperitoneal administration of NF-kappaB p65 siRNA and paclitaxel inhibited cancer growth in mice with peritoneal metastasis of gastric cancer. Therefore, intraperitoneal administration of NF-kappaB p65 siRNA and paclitaxel may provide a breakthrough in the treatment of peritoneal metastasis of gastric cancer.

Elevated Fas expression is related to increased apoptosis of circulating CD8+ T cell in patients with gastric cancer.

BACKGROUND: Extensive apoptosis of immune cells occurs in patients with cancer, and is possibly related to immune evasion by cancer cells. The present study was designed to investigate the correlation between apoptosis levels and Fas expression in CD8+ T lymphocytes in patients with gastric cancer. METHODS: The expression of apoptosis markers (annexin V binding and caspase-3 activation) and the death receptor Fas in CD8+ T cells was evaluated by multicolor flow cytometry. Soluble Fas ligand (sFasL) in the sera was quantitated by enzyme-linked immunosorbent assay. RESULTS: In patients with gastric cancer, 18.7% +/- 10.5% (mean +/- SD) of CD8+ T cells bound annexin V compared with 11.7% +/- 7.9% in normal controls (P = 0.0282). Fas expression in CD8+ T cells was higher in patients with gastric cancer (69.2% +/- 15.3%) than normal controls (50.6% +/- 15.3%) (P = 0.0051). The proportion of apoptotic CD8+ T cells was significantly correlated with Fas expression in CD8+ T cells (r = 0.409, P = 0.0214). In patients, Fas+CD8+ T cells preferentially underwent apoptosis and showed high caspase-3 activation. Moreover, the proportion of apoptotic CD8+ T cells was inversely correlated with serum levels of soluble Fas ligand (r = -0.324, P = 0.0359). Fas expression in tumor infiltrating CD8+ T cells was significantly more frequent (80.3% +/- 13.4%) than in circulating CD8+ T cells (67.9% +/- 15.5%) (P = 0.0046). A decrease in the percentage of Fas+CD8+ T cells was observed after surgery (54.1% +/- 12.8%) compared with before surgery (65.9% +/- 17.0%) (P = 0.0284). CONCLUSIONS: The data indicate that up-regulation of Fas expression in CD8+ T cells is related to increased apoptosis of circulating CD8+ T cells in patients with gastric cancer. Further investigations into the detailed mechanism of apoptosis induction in Fas-positive CD8+ T cells are urgently required.

Decreased NKG2D expression on CD8+ T cell is involved in immune evasion in patients with gastric cancer.

PURPOSE: Some studies suggest that the immunoreceptor NKG2D expression on CD8(+) T cells is down-regulated and this reduction may be involved in immune evasion in cancer patients. The present study was designed to investigate NKG2D expression on CD8(+) T lymphocytes and its relationship to immune evasion in gastric cancer patients. EXPERIMENTAL DESIGN: NKG2D expression on both circulating and tumor-infiltrating CD8(+) T cells was evaluated by multicolor flow cytometry. Soluble MHC class I chain-related gene A (MICA) in the sera was quantitated by ELISA. Transwell experiments were carried out to determine the effect of cancer cells on NKG2D expression. RESULTS: NKG2D expression on circulating CD8(+) T cells was down-regulated and significantly correlated with IFN-gamma production in gastric cancer patients (r = 0.68; P = 0.007). NKG2D expression was closely related to undifferentiated cancer (P = 0.021) as was the depth of invasion (P = 0.012). There was no difference in soluble MICA between gastric cancer patients and normal controls. NKG2D expression on CD8(+) T cells was remarkably reduced in the tissue of gastric cancer compared with peripheral blood (P = 0.046). Complete removal of tumor by surgery restored NKG2D expression on CD8(+) T cells (P = 0.0049). Transwell experiments showed that this down-regulation was induced by direct contact between cancer cells and CD8(+) T cells and that soluble factors did not affect the NKG2D expression. This phenomenon was blocked by the addition of anti-MICA antibodies. CONCLUSIONS: Decreased NKG2D expression may be one of the key mechanisms responsible for immune evasion by tumors in gastric cancer.

Chelation of cadmium ions by phytochelatin synthase: role of the cysteine-rich C-terminal.

The interactions between Cd(2+) and the C-terminal region of phytochelatin (PC) synthase using recombinant wild-type and mutant PC synthase were studied. We show that site-directed mutagenesis of Cys residues at C(358)C(359)XXXC(363)XXC(366) motif decreases the number of Cd(2+) and other heavy metal ions interacting with the enzyme, and that the motif binds the metals discriminatingly. The optimum binding ratio of PC synthase to Cd(2+) was also determined. The findings indicate that Cys exists as a free SH residue and that it is involved in the regulation of PC enzyme activity by transferring the metals into closer proximity with the catalytic domain. These results are important in understanding heavy metal detoxification mechanisms in higher plants, a step towards phytoremediated-applications.