Nonviable Burkholderia mallei induces a mixed Th1- and Th2-like cytokine response in BALB/c mice

Nonviable cell preparations of Burkholderia mallei, the causative agent of glanders, were evaluated as potential vaccine candidates in a BALB/c murine model. Three different B. mallei cell preparations plus Alhydrogel were evaluated: a heat-killed preparation, an irradiation-inactivated preparation, and a preparation of a capsule-negative mutant strain which had been irradiation inactivated. BALB/c mice were vaccinated twice with the different B. mallei preparations, and spleens and sera were collected to determine their cellular and humoral immune responses. All three bacterial cell preparations had essentially the same results in two cellular immune response assays. In a splenocyte proliferation assay, the amount of cell proliferation in response to the homologous immunogen, concanavalin A, or lipopolysaccharide was similar for all the cell preparations. Also, splenocytes from the inoculated mice expressed interleukin 2 (IL-2), gamma interferon, and small amounts of IL-4 and IL-5, and more IL-10 cytokine in the presence of the homologous antigen. When the immunoglobulin subclasses from these mice were examined, they all produced higher levels of IgG1 than IgG2a subclasses. The higher ratio of IgG1 to IgG2a was not due to the amount of the immunogen or the adjuvant (Alhydrogel) used in the BALB/c mice. The cell preparations did not protect the vaccinated mice from a live challenge (>300 50% lethal doses). Our results suggest that in BALB/c mice, a mixed T-helper-cell-like response to nonviable B. mallei is obtained, as demonstrated by a Th1- and Th2-like cytokine response and a Th2-like subclass immunoglobulin response. This may be the reason for the inability of the B. mallei cells that were examined as candidate vaccines to protect the mice from a live challenge.     

Notch-RBP-J signaling is involved in cell fate determination of marginal zone B cells

RBP-J is a key mediator of Notch signaling that regulates cell fate determination in various lineages. To investigate the function of Notch-RBP-J in mature B cell differentiation, we generated mice that selectively lacked B cell RBP-J expression using conditional mutagenesis. Absence of RBP-J led to the loss of marginal zone B (MZB) cells with a concomitant increase in follicular B cells; in contrast, B1 cells in the peritoneal cavity were unaffected. Lack of RBP-J caused no defects in B cells maintenance, survival, plasma cell differentiation or activation. It is therefore likely that Notch-RBP-J signaling regulates the lineage commitment of mature B cells into follicular versus MZB cells. In addition, in mice with RBP-J-deficient B cells, had no obvious changes in immunoglobulin production in response to Ficoll, lipopolysaccharide or chicken gammaglobulin. In contrast, these mice exhibited increased mortality rates after blood-borne bacterial infection, which indicates that MZB cells play pivotal roles in the clearance of these bacteria.     

Granulocytosis associated with malignant neoplasms: a clinicopathologic study and demonstration of colony-stimulating activity in tumor extracts

We studied seven cases of malignant neoplasms, taken from various sites in the body, that were associated with marked granulocytosis. The seven cases were characterized clinically by marked granulocytosis with mature neutrophils, nonspecific inflammatory signs, and a rapid and progressive fatal course of the disease. The elevation of granulocyte count generally paralleled the increase in tumor size. Postmortem examination revealed no evidence of extensive bone marrow metastases or significant suppuration in any case. The bone marrows showed varying degrees of granulocytic hyperplasia with or without a shift to the left in the maturation series. Erythroid cell hyperplasia was observed in some cases, and in one instance there was an increase in immature eosinophils. The spleen showed various degrees of infiltration by neutrophils, from minimal to extremely marked; some spleens had foci of extramedullary hemopoiesis. Colony-stimulating activity was demonstrated in tumor extracts from three of these cases and from the serum in another case. Thus, it is suggested that marked granulocytosis in these patients was caused, at least in part, by colony-stimulating factor produced by the neoplastic cells.     

Nuclear magnetic resonance studies on polymer carbanions, 5. Polystyryl carbanion with lithium as counter cation and its dimer model compounds

Oligostyryllithium was synthesized by the reaction of sec-butyllithium with styrene in diethyl ether at ?40°C. Dimer model anions (counter cations: Li and K) of the growing chain ends were synthesized by metallation of 1-methoxy-1,3-diphenylpropane. The structures of the lithium compounds were corroborated by ¹³C NMR spectroscopy and compared with those of the potassium compounds. As for polystyryllithium and its model compounds, the charge density on the α-carbon increased and that on the phenyl ring of the growing end decreased with increasing molecular weight, while, in polystyrylpotassium and its model compounds, the charge density on the α-carbon increased and the one on the phenyl ring did not change with increasing molecular weight. The temperature change of the ¹H NMR spectrum of oligostyryllithium in diethyl ether indicated rotation around the C-1? C_α bond at lower temperatures than in tetrahydrofuran, but the activation energy was the same in both solvents. T₁ values of the phenyl carbons in model compound 2b were one to two orders of magnitude shorter than those of the protonated compound.     

Immunohistochemical analysis of centromere protein F expression in buccal and gingival squamous cell carcinoma

Centromere protein F (CENP-F) expression (localization and characteristics) in relation to tumor clinicopathological parameters was immunohistochemically examined and evaluated in 47 archival biopsy specimens of buccal and gingival squamous cell carcinomas (SCC). Centromere protein F expression was detected in 79% of the samples. An increase in the labeling index (LI) with WHO grading was obtained ( P  < 0.05). Correlations were obtained between the CENP-F LI and tumor size ( P  < 0.05). Immunoelectron microscopy showed CENP-F nuclear staining as punctate or fine dots. The present study shows that CENP-F expression and detection of a more specific cell subpopulation presents a theoretical advantage for the analysis of the precise cell cycle of G₂ to M cells, compared to Ki-67.     

Minimally immunogenic decellularized porcine valve provides in situ recellularization as a stentless bioprosthetic valve

Currently used bioprosthetic valves have several limitations such as calcification and functional deterioration, and revitalization through cellular ingrowth is impossible. To overcome these obstacles, we have developed a minimally immunogenic tissue-engineered valve that consists of an unfixed, decellularized porcine valve scaffold capable of being spontaneously revitalized in vivo after implantation. Porcine aortic root tissue was decellularized using detergents such as sodium lauryl sulfate and Triton X-100. The porcine valve was treated very gently and plenty of time was allowed for constituents to diffuse in and out of the matrix. In a preliminary study, a piece of decellularized porcine valve tissue was implanted into the rat subdermal space for 14 and 60 days and the structural integrity and calcification were evaluated. As an in vivo valve replacement model, the decellularized porcine valve was implanted in the pulmonary valve position in dogs and functional and histological evaluation was performed after 1, 2, and 6 months. Histological examination showed that the newly developed detergent treatment effectively removed cellular debris from the porcine aortic tissue. Decellularized porcine valve tissue implanted subdermally in rats showed minimal inflammatory cell infiltration and calcification. In the valve replacement model, spontaneous reendothelialization and repopulation of the medial cells were observed within 2 months, and good valve function without regurgitation was observed by echocardiography up to 6 months. The minimally immunogenic decellularized porcine valve proved effective in mitigating postimplant calcification and provided a suitable matrix for revitalizing prostheses through in situ recellularization, cellular ingrowth, and tissue remodeling.     

Investigation of albumin synthesis in rat livers during the perinatal period using immunocytochemistry andin situ hybridization

The immunoreactivity of albumin (ALB) was observed in the hepatocytes of fetal rats on day 18 of gestation, and was especially observable in immature rough endoplasmic reticulum (rER) and Golgi apparatus (GA); by then, a small amount of silver grains of ALB mRNA could already be detected. Just after birth, immunoreactivity of ALB could be observed in fine granules or diffusely in all hepatocytes, and was present in rER and GA. One week after birth immunoreactivity of ALB was observed in all hepatocytes and was visible in developed rER and GA; the grains of ALB mRNA were present in all hepatocytes.     

Clinicopathological significant and prognostic influence of cadherin-17 expression in gastric cancer

Cadherin-17 (CDH17), also called liver–intestine cadherin, is a structurally unique member of the cadherin superfamily. Our serial analysis of gene expression demonstrated that CDH17 was one of the most up-regulated genes in advanced gastric carcinomas. CDH17 expression is known to be regulated by Cdx2. In the present study, we examined the expression of CDH17 in primary gastric carcinoma tissues by immunohistochemistry, and analyzed the correlation of CDH17 expression with clinicopathological characteristics and patients prognosis. CDH17 expression was detected in 63/94 (67%) of gastric adenocarcinomas in addition to intestinal metaplasia. The expression of CDH17 tended to be associated with intestinal type carcinoma, and carcinomas with CDH17 expression was significantly more frequent in advanced stage cases (80%) than in early stage (53%). The prognosis of patients with positive CDH17 expression was significantly poorer than that of the negative cases (P=0.0314). However, multivariate analysis revealed that CDH17 was not an independent prognostic factor. Six of seven cases that showed positive expression of Cdx2 simultaneously expressed CDH17 protein. These results suggested that the expression of CDH17 was characteristic of the advanced gastric carcinoma that is associated with poor prognosis.     

ABC proteins: key molecules for lipid homeostasis

Forty-nine ABC protein genes exist on human chromosomes. Eukaryotic ABC proteins were originally recognized as drug efflux pumps involved in the multidrug resistance of cancer cells. However, it is now realized that one of their major physiological roles is cellular lipid transport and homeostasis, and their dysfunction is often associated with human diseases. ABCA1 and ABCA7 mediate the apolipoprotein-dependent formation of a high-density lipoprotein–cholesterol complex. ABCA3 is indispensable for pulmonary surfactant secretion. ABCG5 and ABCG8 are involved in the secretion of plant sterols and cholesterol into bile. However, the primary substrates and mechanism of action of these ABC proteins have not been precisely defined. In this review article, we first describe the general structure and functions of eukaryotic ABC proteins. The current model of ABCA1 functionality is then explained based on studies on a topological model, subcellular localization, apoA-I dependence of HDL formation, functional defects of Tangier disease mutants, and ATP hydrolysis of purified ABCA1. ABCA1 is supposed to function as a transporter of lipids as well as a receptor for apoA-I. ABCA3 is likely involved in accumulating phospholipids and cholesterol in lamellar bodies and in generating multivesicular structures.     

Autoimmunity and inflammation due to a gain-of-function mutation in phospholipase C gamma 2 that specifically increases external Ca2+ entry

The identification of specific genetic loci that contribute to inflammatory and autoimmune diseases has proved difficult due to the contribution of multiple interacting genes, the inherent genetic heterogeneity present in human populations, and a lack of new mouse mutants. By using N-ethyl-N-nitrosourea (ENU) mutagenesis to discover new immune regulators, we identified a point mutation in the murine phospholipase Cg2 (Plcg2) gene that leads to severe spontaneous inflammation and autoimmunity. The disease is composed of an autoimmune component mediated by autoantibody immune complexes and B and T cell independent inflammation. The underlying mechanism is a gain-of-function mutation in Plcg2, which leads to hyperreactive external calcium entry in B cells and expansion of innate inflammatory cells. This mutant identifies Plcg2 as a key regulator in an autoimmune and inflammatory disease mediated by B cells and non-B, non-T haematopoietic cells and emphasizes that by distinct genetic modulation, a single point mutation can lead to a complex immunological phenotype.