Molecular genetic analysis of Actinobacillus actinomycetemcomitans epidemiology

Research over the past decade has identified many of the microorganisms involved in the etiology of human periodontitis such as Actinobacillus actinomycetemcomitans. Efforts are now directed toward defining these species' role in the pathogenic process. Since microbial colonization of host tissues is a key first step in developing a bacterial infection, determining the source of the periodontal pathogens and their route of transmission is likely to be crucial in formulating preventive strategies. Recently, a technique from molecular biology, restriction endonuclease analysis, has been used to track bacterial infections. In the present study, this method was used to investigate the epidemiology of A. actinomycetemcomitans infection. One hundred twenty-four human subgingival plaque isolates of A. actinomycetemcomitans were examined including bacterial strains from the United States, Korea, and Norway as well as 15 strains from cynomolgus (Macaca fascicularis) and spider monkeys (Macaca iris) and 4 reference strains. The genomic DNA from each strain was purified, digested with each of 16 restriction endonucleases, and the DNA digests were resolved by electrophoresis. The resulting patterns of DNA fragments were compared and also correlated with the A. actinomycetemcomitans serotype determined using serotype-specific antisera in immunofluorescence. Human isolates of A. actinomycetemcomitans even from disparate geographic sources showed little diversity by restriction endonuclease analysis. Three major restriction patterns were found. Restriction pattern I was common to all 20 of the serotype a isolates, restriction pattern II was associated with 58% of the 73 serotype b isolates examined, while restriction pattern III was associated with the remaining serotype b strains and with all 15 of the serotype c strains.(ABSTRACT TRUNCATED AT 250 WORDS)     

National Study of the Relation of Primary Care Shortages to Emergency Department Utilization

Background: Emergency department (ED) visit volumes are increasing nationwide. Objectives: To determine whether states with primary care shortages have higher rates of ED use. Methods: Populations residing in primary care shortage areas were abstracted from the Health Resources and Services Administration Geospatial Database. Annual ED visit volumes were available from the 2001 National ED Inventory. Population data and potential confounders were abstracted from federal data sets. All analyses were conducted at the state level. Results: Primary care shortage densities varied greatly across states, ranging from 3 (New Jersey) to 28 (Mississippi) medically underserved individuals per 100 people. States also varied in their annual ED visit densities, ranging from 23 visits (Hawaii) to 65 visits (Washington, DC) per 100 people. Of the 17 states in the top tertile for primary care shortage, 7 also were in the top tertile for ED visits. Primary care shortage density was positively associated with ED visit density. An increase of 10 medically underserved individuals per 100 people was associated with an annual increase of 4.2 ED visits per 100 people (p = 0.04). The association remained after controlling for six factors, with an increase of 10 medically underserved individuals per 100 people associated with an annual increase of 3.3 ED visits per 100 people (p = 0.04). Nevertheless, five states had high ED visit densities despite comparatively low primary care shortage densities (Maine, Massachusetts, New Hampshire, Rhode Island, and Vermont), whereas five others had low ED visit densities despite high primary care shortage densities (Arizona, Idaho, Montana, New Mexico, and South Dakota). Conclusions: A positive association between primary care shortage densities and ED visit densities was found. Although most states adhere to this pattern, some states do not. Further investigation of this dissociation may yield additional explanations for rising ED visit volumes.     

Metabolism of a glucuronide conjugate of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone in rats

 Besides 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), [4-(methylnitrosamino)-1-(3-pyridyl)but-1-yl]-β-O-d-glucosiduronic acid (NNAL-Glu) is another important metabolite of the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) which has been detected in the urine of tobacco users and non-smokers heavily exposed to sidestream cigarette smoke. In order to evaluate the toxicological significance of NNAL-Glu formation and excretion, the metabolism of [5-³H]-NNAL-Glu was studied in rats. Five male F344 rats were administered 3.7 mg/kg [5-³H]-NNAL-Glu by i.v. injection and the metabolites in urine analysed by HPLC. More than 90% of the radioactivity was excreted in urine within the first 24 h. Unchanged NNAL-Glu accounted for 81.2±3.1% of the total radioactivity; the remaining part of the dose appears to be deconjugated resulting in the urinary excretion of NNAL (3.6±1.7%) and its α-hydroxylation (11.5±2.2%) and N-oxidation (3.6±1.6%) products. The presence of α-hydroxylation products of NNAL-Glu in urine suggests that this NNK metabolite may be activated in vivo to carcinogenic intermediates. Received: 25 April 1994 / Accepted: 29 June 1994     

Plasma concentrations of C-reactive protein and total homocysteine in relation to the severity and risk factors for cerebrovascular disease

Higher C-reactive protein (CRP) and plasma homocysteine (tHcy) concentrations have been shown to indicate increased risk of coronary heart disease and cerebrovascular disease (CVD), but the mechanisms by which they increase the risk of atherothrombotic disease are under investigation. This study evaluates the associations of high-sensitivity C-reactive protein (hs-CRP) and tHcy with the risk factors, severity, and outcome on discharge in patients with CVD. hs-CRP, fasting tHcy, and lipid profile were determined in 50 patients with CVD and 20 healthy control subjects. Clinical data, National Institutes of Health stroke scale (NIHSS) on admission and disability Rankin scale on discharge, were recorded. Based on epidemiologic studies, cutoff points of 1.5 mg/L (hs-CRP) and 15mumol/L (tHcy) were used to indicate increased risk. Univariate and multivariate logistic regression analyses were used to relate tHcy with other CVD risk factors, NIHSS on admission and the disability Rankin scale on discharge. Overall, 38% of patients had increased hs-CRP and 26% had elevated tHcy. hs-CRP (P = 0.005) and tHcy (P < 0.0001) concentrations were significantly higher in patients compared with controls, and these differences remained significant after correction for age and sex. tHcy showed significant correlations with hs-CRP (rs = 0.35; P = 0.003) and low-density lipoprotein-cholesterol (LDL-C; rs = 0.49; P = 0.005). Logistic regression analysis with CVD as the dependent variable showed significant association with hs-CRP (P = 0.01) and tHcy (P < 0.0001) after adjustment for potential confounders. hs-CRP showed increased trend with disease severity and significant association with the disability Rankin scale (P = 0.033). These data support 4 main conclusions: (1) Elevation of hs-CRP and tHcy are common in CVD; (2) the significant relationship between tHcy and hs-CRP suggests that the association of tHcy with CVD risk may be dependent on inflammation-related mechanisms; (3) increased hs-CRP and tHcy show that patients with CVD may be at greater risk of subsequent coronary heart disease; and (4) admission hs-CRP could be used as an indicator of prognosis.     

Association of high-sensitivity C-reactive protein and odds of breast cancer by molecular subtype: analysis of the MEND study

Breast cancer (BC) in Nigeria is characterized by disproportionately aggressive molecular subtypes. C-reactive protein (CRP) is associated with risk and aggressiveness for several types of cancer. We examined the association of high-sensitivity CRP (hsCRP) with odds of BC by molecular subtype among Nigerian women. Among 296 newly diagnosed BC cases and 259 healthy controls, multivariable logistic regression models were used to estimate adjusted odds ratios (aOR) and 95% confidence intervals (CI) for the association between hsCRP and odds of BC overall and by molecular subtype (luminal A, luminal B, HER2-enriched and triple-negative or TNBC). High hsCRP (> 3 mg/L) was observed in 57% of cases and 31% of controls and was associated with 4 times the odds of BC (aOR: 4.43; 95% CI: 2.56, 7.66) after adjusting for socio-demographic, reproductive, and clinical variables. This association persisted regardless of menopausal status and body mass index (BMI) category. High hsCRP was associated with increased odds of TNBC (aOR: 3.32; 95% CI: 1.07, 10.35), luminal A BC (aOR: 4.03; 95% CI: 1.29, 12.64), and HER2-enriched BC (aOR: 6.27; 95% CI: 1.69, 23.25). Future studies are necessary in this population to further evaluate a potential role for CRP as a predictive biomarker for BC.     

Upregulation of CD4+ T-Lymphocytes by Isomeric Mixture of Quercetin-3-O-Rutinoside and Quercetin-3-O-Robinobioside Isolated from Millettia aboensis

Background: Millettia aboensis (Hook. F.) Baker (Fabaceae) is popular in ethnomedicine for its acclaimed efficacy in a number of disease conditions. This study evaluated the immunomodulatory effect of the leaf extract as a possible mechanism of its ethnomedicinal uses.

Methods: Humoral and cellular immune responses of Balb/c mice to tetanus toxoid and cyclophosphamide, respectively, were used to monitor immunomodulatory activities of the ethanol leaf extract and fractions of M. aboensis at 200, 300 and 400 mg/kg. Active (butanol) fraction of the extract was subjected to chromatographic purifications to isolate the active compound and the structure elucidated by a combination of 1D and 2D NMR and mass spectrometry. Stimulation of specific T-lymphocytes using intracellular cytokine staining technique was used to evaluate immune-enhancing activity of the isolated compound.

Results: The extract and fractions evoked increase in both humoral and cellular immunity. At 400 mg/kg of butanol fraction, the normalized mean secondary production of IgG1 and IgG2a antibodies were 9.0 and 7.7, respectively. Serum cytokine production by butanol fraction following secondary challenge with tetanus toxoid showed that IL-12, IL-17A and IFN-γ were expressed by 48.14, 41.37 and 38.22%, respectively. Structural elucidation of the active compound revealed presence of isomeric mixtures of quercetin-3-O-rutinoside and quercetin-3-O-robinobioside (Compound 1a/b). Compound 1a/b exhibited in vitro upregulation of specific CD4⁺ T-lymphocytes that were largely IFNγ releasing with up to 43.7% stimulation at 6.25 μg/mL compared to the baseline effect in DMSO vehicle control group.

Conclusion: M. aboensis expressed strong immune-enhancing properties, which may explain its ethnopharmacological use in disease management.     

Differential screening of a human chromosome 3 library identifies hepatocyte growth factor-like/macrophage-stimulating protein and its receptor in injured lung. Possible implications for neuroendocrine cell survival.

Transient pulmonary neuroendocrine cell hyperplasia and non-neuroendocrine lung tumors develop in nitrosaminetreated hamsters, which we hypothesized might modulate epithelial cell phenotype by expressing gene(s) homologous to human chromosome 3p gene(s) deleted in small cell carcinoma of the lung (SCLC). We differentially screened a chromosome 3 library using nitrosamine-treated versus normal hamster lung cDNAs and identified hepatocyte growth factor-like/macrophage-stimulating protein (HGFL/MSP) in injured lung. HGFL/MSP mRNA is low to undetectable in human SCLC and carcinoid tumors, but the HGFL/MSP tyrosine kinase receptor, RON, is present and functional on many of these neuroendocrine tumors. In H835, a pulmonary carcinoid cell line, and H187, a SCLC cell line, HGFL/ MSP induced adhesion/flattening and apoptosis. Using viable cell counts to assess proliferation after 14 d of treatment with HGFL/MSP, there is growth inhibition of H835 but not H187. Nitrosamine-treated hamsters also demonstrate pulmonary neuroendocrine cell apoptosis in situ during the same time period as expression of the endogenous HGFL/ MSP gene, immediately preceding the spontaneous regression of neuroendocrine cell hyperplasia. These observations suggest that HGFL/MSP might regulate neuroendocrine cell survival during preneoplastic lung injury, which could influence the ultimate tumor cell phenotype.