BCL2 translocations in leukemias of mature B cells

Although translocations of the BCL2 gene are frequent in B-cell non- Hodgkin's lymphomas (B-NHL) the incidence, nature, and prognostic significance of similar translocations in the phenotypically related chronic leukemias of mature B cells are unknown. Therefore, we examined 170 cases of B-cell chronic lymphocytic leukemia (B-CLL), 7 cases of B- cell prolymphocytic leukemia (B-PLL), 25 cases of hairy cell leukemia (HCL) and 22 cases of splenic lymphoma with villous lymphocytes (SLVL) with defined cytogenetic abnormalities by DNA blot using both 5' and 3' BCL2 probes to search for rearrangement of the BCL2 locus. Translocation t(14;18) (q32.3;q21.3) was detected cytogenetically in 3 cases of B-CLL. All had breakpoints in the 3' region of BCL2, mapping between the major breakpoint region (MBR) and the minor cluster region (mcr), the breakpoint clusters commonly detected in B-NHL. In 2 of the 3 cases, the breakpoint within BCL2 was mapped to a 1.0-kb EcoRI- HindIII fragment indicating a clustering of breakpoints. Two cases of B- CLL had cytogenetically detectable t(2;18)(p11;q21.3) or t(18;22)(q21.3;q11). Both had rearranged the 5' region of the BCL2 gene to the corresponding lg light-chain gene. Molecular cloning of the t(18;22)(q21.3;q11) showed that the translocation disrupted the BCL2 promoter region and the first untranslated BCL2 exon. Nevertheless, high levels of BCL2 protein were seen in this case. Only 2 other cases in whom cytogenetic analysis was not successful showed rearrangement of the 5' region of BCL2, an overall incidence of 2.3%. No cases of B-PLL, HCL, or SLVL showed either 5' or 3' BCL2 rearrangement. These data confirm the cytogenetic observations that translocations involving the BCL2 locus in all forms of leukemia of mature B cells are rare, and limited to a minor subset of B-CLL. BCL2 translocations in B-CLL involve hot spots of recombination of both the 5' and 3' regions of the BCL2 gene, which are distinct from those commonly seen in B-NHL, suggesting distinct pathogenic mechanisms.     

Speed of healing and symptom relief in grade II to IV gastroesophageal reflux disease: A meta-analysis

BACKGROUND & AIMS: Esophagitis healing proportions are often incorrectly called the healing rate. The aim of this study was to compare different drug classes by expressing the speed of healing and symptom relief through a new approach. METHODS: A fully recursive literature search to July 1996 identified 43 articles on gastroesophageal reflux disease (GERD) (7635 patients) meeting strict inclusion criteria: single- or double-blind randomized studies in adults with endoscopically proven erosive or ulcerative esophagitis. For each drug class, linear regression analysis estimated the average percentage of patients who were healed and heartburn free per week. RESULTS: Mean overall healing proportion irrespective of drug dose or treatment duration (< or =12 weeks) was highest with proton pump inhibitors (PPIs; 83.6% +/- 11.4%) vs. H2-receptor antagonists (H2RAs; 51.9% +/- 17.1%), sucralfate (39.2% +/- 22.4%), or placebo (28.2% +/- 15.6%). Correcting for patients without baseline heartburn, the mean heartburn-free proportion was highest with PPIs (77.4% +/- 10.4%) vs. H2RAs (47.6% +/- 15.5%). PPIs showed a significantly faster healing rate (11.7%/wk) vs. H2RAs (5.9%/wk) and placebo (2.9%/wk). PPIs provided faster, more complete heartburn relief (11.5%/wk) vs. H2RAs (6.4%/wk). CONCLUSIONS: More complete esophagitis healing and heartburn relief is observed with PPIs vs. H2RAs and occurs nearly twice as fast. This semiquantitative expression of speed of healing and symptom relief permits comparisons for future economic evaluation and quality-of-life assessments. (Gastroenterology 1997 Jun;112(6):1798-810)     

Chronic kinin receptor blockade attenuates the antihypertensive effect of ramipril.

The contribution of endogenous kinins to the chronic antihypertensive effect of angiotensin converting enzyme inhibitors was investigated in two-kidney, one clip hypertensive Wistar rats, using the new bradykinin B2-receptor antagonist HOE 140 (D-Arg, [Hyp3, Thi5, D-Tic7, Oic8]-bradykinin). In a first protocol, rats were pretreated orally with the angiotensin converting enzyme inhibitor ramipril (1 mg/kg per day), for 4 weeks. Acute blockade of bradykinin receptors by intravenous injections of HOE 140 at doses of 8.4 and 100 micrograms/kg, which inhibited the depressor responses to exogenous bradykinin, did not affect the antihypertensive effect of ramipril in these animals. Bradykinin receptors were then blocked chronically by subcutaneous infusion of HOE 140 (500 micrograms/kg per day) via osmotic minipumps for 6 weeks, while ramipril treatment was continued. HOE 140 partially reversed the antihypertensive effect of ramipril from 115.3 +/- 4.6 to 123.8 +/- 3.3 mm Hg (mean arterial blood pressure) after 3 weeks and to 121.3 +/- 2.9 mm Hg after 6 weeks. In contrast, in controls (ramipril plus subcutaneous vehicle infusion) mean arterial blood pressure decreased further from 112.0 +/- 6.0 to 110.3 +/- 4.9 mm Hg after 3 weeks and to 103.7 +/- 5.0 mm Hg after 6 weeks (p less than 0.05 and p less than 0.01, HOE 140 versus controls). Plasma catecholamines were not significantly different between the two groups at the end of the experiment, indicating that the partial reversal of the antihypertensive effect was not due to a bradykinin-like agonistic effect on catecholamine release.(ABSTRACT TRUNCATED AT 250 WORDS)     

Calin from Hirudo medicinalis, an inhibitor of platelet adhesion to collagen, prevents platelet-rich thrombosis in hamsters

Interaction between exposed collagen and platelets and/or von Willebrand factor is believed to be one of the initiating events for thrombus formation at sites of damaged endothelium. Interference with this mechanism may provide an anti-thrombotic potential. Calin, a product from the saliva of the leech Hirudo medicinalis, was tested in vitro and for its in vivo activity in a thrombosis model in hamsters. Calin specifically and dose dependently (IC50:6.5 to 13 micrograms/mL) inhibited human platelet aggregation induced by collagen. In addition, specific platelet adhesion onto microtiter wells coated with collagen and detected with a monoclonal antiglycoprotein IIb/IIIa antibody- conjugated with horseradish peroxidase, could be completely prevented with Calin (IC50:22 micrograms/mL). A dose-response curve was constructed in groups of six hamsters in whom a standardized trauma was induced on the femoral vein. Thrombus formation was followed continuously using video recording and processing of the image obtained upon transillumination of the vessel. Intravenous Calin dose- dependently inhibited platelet-rich thrombus formation in this model with an ED50 of 0.07 mg/kg and complete inhibition with 0.2 mg/kg. No effects were seen on coagulation tests or bleeding times, whereas ex vivo aggregation induced by collagen was inhibited dose dependently. Local application of leech saliva, Calin, hirudin, or the combination of the latter two into the bleeding time wound of hamsters resulted in a mild prolongation of the bleeding time (twofold to threefold). A similar experiment in baboons did not cause any prolongation of the bleeding time. This is in sharp contrast with the long-lasting bleeding after a leech bite itself in both species. Calin from the leech Hirudo medicinalis is able, by binding to collagen, to effectively interfere with platelet-collagen interaction, which results in an antithrombotic effect observed in a platelet-rich thrombosis model in hamsters.     

Protease‐activated receptor 2 suppresses lymphangiogenesis and subsequent lymph node metastasis in a murine pancreatic cancer model

Protease‐activated receptor‐2 (PAR‐2) is a G protein‐coupled receptor that functions as a cell‐surface sensor for coagulation factors and other proteases associated with the tumour microenvironment. Pancreatic cancer cells express high levels of PAR‐2 and activation of PAR‐2 may induce their proliferation and migration. Interestingly, however, PAR‐2 expression is increased in stroma‐rich pancreatic cancer regions, suggesting a potential role of PAR‐2 in the tumour microenvironment. Here, we assessed the importance of PAR‐2 in the stromal compartment by utilizing an orthotopic pancreatic cancer model, in which tumour cells are PAR‐2‐positive, whereas stromal cells are PAR‐2‐negative. We assessed tumour weight and volume and analysed proliferation and (lymph)angiogenesis both in vivo and in vitro. We show that genetic ablation of PAR‐2 from the stromal compartment inhibits primary tumour growth, which is accompanied by reduced vascularization in primary tumours and reduced in tube formation of vascular endothelial cells in vitro. In contrast to smaller primary tumours, the number of lymph node metastases was increased in PAR‐2‐deficient animals, which was accompanied by an increased number of lymphatic vessels. In vitro tube‐formation assays show that PAR‐2 does not inhibit the intrinsic tube‐forming capacity of lymphatic endothelial cells, but that PAR‐2 actually inhibits cancer cell‐induced tube formation. Overall, stromal PAR‐2 thus plays a dual role in pancreatic cancer development by potentiating primary tumour growth but limiting lymphangiogenesis and subsequent lymph node metastasis. Our data identify a novel role of PAR‐2 in the tumour microenvironment and pinpoint PAR‐2 as a negative regulator of lymphangiogenesis. Copyright © 2014 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Oral health of Adelaide nursing home residents: longitudinal study

Objective: The Adelaide Dental Study of Nursing Homes aimed to quantify oral disease experience, incidence and increments in Adelaide nursing home residents. Methods: Questionnaires and dental inspections were completed at baseline and at 1‐year for residents from randomly selected Adelaide nursing homes. Results: The residents were very functionally dependent, cognitively impaired and behaviourally difficult older adults with complex oral problems and dental treatment needs. The prevalence of edentulism (total tooth loss) (63%) decreased and more residents were retaining natural teeth. Existing residents had a mean of 10.8 teeth present and new residents had a mean of 12.7 teeth present. Residents’ previous experiences of caries (decay) were high – existing residents had a mean of 1.2 decayed teeth and new residents had a mean of 0.8 decayed teeth. Residents’ caries increments (new decay) over the 1‐year period were high (coronal = 2.5 surfaces; root = 1.0 surfaces), especially in those who had lost weight and who could eat fewer food types. These levels of caries were many times greater than had been reported for community‐dwelling older adults. Large accumulations of plaque, calculus and debris (food) were evident on residents’ natural teeth and dentures, especially those with dementia. Up to 25% of residents owned dentures that were not worn. Residents with dementia gave their carers complex and challenging oral hygiene care problems. Existing and new residents had similar general health and oral health characteristics, with the exceptions that new residents had significantly more filled tooth surfaces, and fewer decayed retained roots. Conclusion: New residents were being admitted to the nursing homes with a compromised oral health status or developed severe oral diseases and conditions within several months of their admittance. Residents’ oral diseases, especially coronal and root caries, rapidly progressed during their stay in residential care.     

Insulin‐like growth factor‐II is produced by,signals to and is an important survival factor for the mature podocyte in man and mouse

Podocytes are crucial for preventing the passage of albumin into the urine and, when lost, are associated with the development of albuminuria, renal failure and cardiovascular disease. Podocytes have limited capacity to regenerate, therefore pro‐survival mechanisms are critically important. Insulin‐like growth factor‐II (IGF‐II) is a potent survival and growth factor; however, its major function is thought to be in prenatal development, when circulating levels are high. IGF‐II has only previously been reported to continue to be expressed in discrete regions of the brain into adulthood in rodents, with systemic levels being undetectable. Using conditionally immortalized human and ex vivo adult mouse cells of the glomerulus, we demonstrated the podocyte to be the major glomerular source and target of IGF‐II; it signals to this cell via the IGF‐I receptor via the PI3 kinase and MAPK pathways. Functionally, a reduction in IGF signalling causes podocyte cell death in vitro and glomerular disease in vivo in an aged IGF‐II transgenic mouse that produces approximately 60% of IGF‐II due to a lack of the P2 promoter of this gene. Collectively, this work reveals the fundamental importance of IGF‐II in the mature podocyte for glomerular health across mammalian species. Copyright © 2013 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Population bulk segregant mapping uncovers resistance mutations and the mode of action of a chitin synthesis inhibitor in arthropods

Because of its importance to the arthropod exoskeleton, chitin biogenesis is an attractive target for pest control. This point is demonstrated by the economically important benzoylurea compounds that are in wide use as highly specific agents to control insect populations. Nevertheless, the target sites of compounds that inhibit chitin biogenesis have remained elusive, likely preventing the full exploitation of the underlying mode of action in pest management. Here, we show that the acaricide etoxazole inhibits chitin biogenesis in Tetranychus urticae (the two-spotted spider mite), an economically important pest. We then developed a population-level bulk segregant mapping method, based on high-throughput genome sequencing, to identify a locus for monogenic, recessive resistance to etoxazole in a field-collected population. As supported by additional genetic studies, including sequencing across multiple resistant strains and genetic complementation tests, we associated a nonsynonymous mutation in the major T. urticae chitin synthase (CHS1) with resistance. The change is in a C-terminal transmembrane domain of CHS1 in a highly conserved region that may serve a noncatalytic but essential function. Our finding of a target-site resistance mutation in CHS1 shows that at least one highly specific chitin biosynthesis inhibitor acts directly to inhibit chitin synthase. Our work also raises the possibility that other chitin biogenesis inhibitors, such as the benzoylurea compounds, may also act by inhibition of chitin synthases. More generally, our genetic mapping approach should be powerful for high-resolution mapping of simple traits (resistance or otherwise) in arthropods.     

Clinically Distinct Phenotypes of Canavan Disease Correlate with Residual Aspartoacylase Enzyme Activity

We describe 14 patients with 12 novel missense mutations in ASPA, the gene causing Canavan disease (CD). We developed a method to study the effect of these 12 variants on the function of aspartoacylase—the hydrolysis of N‐acetyl‐l ‐aspartic acid (NAA) to aspartate and acetate. The wild‐type ASPA open reading frame (ORF) and the ORFs containing each of the variants were transfected into HEK293 cells. Enzyme activity was determined by incubating cell lysates with NAA and measuring the released aspartic acid by LC–MS/MS. Clinical data were obtained for 11 patients by means of questionnaires. Four patients presented with a non‐typical clinical picture or with the milder form of CD, whereas seven presented with severe CD. The mutations found in the mild patients corresponded to the variants with the highest residual enzyme activities, suggesting that this assay can help evaluate unknown variants found in patients with atypical presentation. We have detected a correlation between clinical presentation, enzyme activity, and genotype for CD.