CXCL1: A new diagnostic biomarker for human tuberculosis discovered using Diversity Outbred mice

More humans have died of tuberculosis (TB) than any other infectious disease and millions still die each year. Experts advocate for blood-based, serum protein biomarkers to help diagnose TB, which afflicts millions of people in high-burden countries. However, the protein biomarker pipeline is small. Here, we used the Diversity Outbred (DO) mouse population to address this gap, identifying five protein biomarker candidates. One protein biomarker, serum CXCL1, met the World Health Organization’s Targeted Product Profile for a triage test to diagnose active TB from latent M.tb infection (LTBI), non-TB lung disease, and normal sera in HIV-negative, adults from South Africa and Vietnam. To find the biomarker candidates, we quantified seven immune cytokines and four inflammatory proteins corresponding to highly expressed genes unique to progressor DO mice. Next, we applied statistical and machine learning methods to the data, i.e., 11 proteins in lungs from 453 infected and 29 non-infected mice. After searching all combinations of five algorithms and 239 protein subsets, validating, and testing the findings on independent data, two combinations accurately diagnosed progressor DO mice: Logistic Regression using MMP8; and Gradient Tree Boosting using a panel of 4: CXCL1, CXCL2, TNF, IL-10. Of those five protein biomarker candidates, two (MMP8 and CXCL1) were crucial for classifying DO mice; were above the limit of detection in most human serum samples; and had not been widely assessed for diagnostic performance in humans before. In patient sera, CXCL1 exceeded the triage diagnostic test criteria (>90% sensitivity; >70% specificity), while MMP8 did not. Using Area Under the Curve analyses, CXCL1 averaged 94.5% sensitivity and 88.8% specificity for active pulmonary TB (ATB) vs LTBI; 90.9% sensitivity and 71.4% specificity for ATB vs non-TB; and 100.0% sensitivity and 98.4% specificity for ATB vs normal sera. Our findings overall show that the DO mouse population can discover diagnostic-quality, serum protein biomarkers of human TB.     

Serum IL-18 levels in patients with type 1 diabetes: relations to metabolic control and microvascular complications

The study was designed to examine serum IL-18 level and its relation to metabolic control parameters and microvascular complications in type 1 diabetes mellitus (DM). Sixty two patients with type 1 DM and 30 healthy individuals were enrolled in the study. Serum IL-18 levels of patients with type 1 DM were significantly increased compared to controls (293.4+/-133.4 vs 211.2+/-63.9 pg/ml, P=0.003). Patients with poor glycemic control had higher levels of IL-18 than patients with well glycemic control (329.9+/-141.0 vs 226.3+/-89.6 pg/ml, P=0.02). There was no significant difference between the serum IL-18 levels of patients with microvascular complications and those of patients without microvascular complications (307.6+/-127.6 vs 293.2+/-145.6 pg/ml, P>0.05). IL-18 correlated positively with HbA(1c) (r=0.32, P=0.01) and postprandial blood glucose (PPBG) (r=0.26, P=0.02); and negatively with HDL-cholesterol (HDL-C) (r=-0.38, P=0.007). By linear regression analysis, PPBG was determined as the most explanatory parameter for the alterations in serum IL-18 levels (P=0.02). High levels of IL-18 in patients with type 1 DM is related to short and long term glycemic control and HDL-C levels but not to microvascular complications.     

Amide derivatives with pyrazole carboxylic acids of 5-amino-1,3,4-thiadiazole 2-sulfonamide as new carbonic anhydrase inhibitors: synthesis and investigation of inhibitory effects

Pyrazole carboxylic acid amides of 5-amino-1,3,4-thiadiazole-2-sulfonamide were synthesized from 4-benzoyl-1,5-diphenyl-1H-pyrazole-3-carbonyl chloride and 4-benzoyl-1-(3-nitrophenyl)-5-phenyl-1H-pyrazole-3-carbonyl chloride. Carbonic anhydrase isoenzymes (hCA-I and hCA-II) were purified from human erythrocyte cells by the affinity chromatography method. The inhibitory effects of 5-amino-1,3,4-thiadiazole-2-sulfonamide 1, acetazolamide 2 and new synthesized amides on these isozymes have been studied in vitro. The I(50) concentrations (the concentration of inhibitor producing a 50% inhibition of CA activity) against hydratase activity ranged from 1.2 to 2.2 nM for hCA-I and from 0.4 to 2 nM for hCA-II. The I(50) values against esterase activity ranged from 1.4 to 8 nM for hCA-I and from 1.3 to 6 nM for hCA-II. The K(i) values were observed between 8.2 x 10(- 5) to 6.2 x 10(- 4) M for hCA-I and between 2.9 x 10(- 4) to 8.2 x 10(- 4) M for hCA-II. The comparison of new synthesized amides to 5-amino-1,3,4-thiadiazole-2-sulfonamide 1, acetazolamide 2 indicated that the new synthesized compounds (18-23) inhibit CA activity more potently than the parent compounds.     

Leptin Does Not Play a Major Role in Platelet Aggregation in Obesity and Leptin Deficiency

Objective: A recent study suggested that high concentrations of leptin enhance platelet aggregations. Therefore, the aim of this study was to investigate whether platelet aggregation is altered in patients with leptin gene mutations compared with obese subjects or controls. Research Methods and Procedures: Four men (one homozygous man and his three heterozygous brothers) carrying a leptin gene mutation; 20 age‐matched, healthy, unrelated men; and 18 age‐matched obese men were enrolled in the study. Adenosine diphosphate (ADP)‐, collagen‐, and epinephrine‐induced platelet aggregation were evaluated in all individuals. Results: Our results show that patients with the leptin gene mutation (both the homozygous and heterozygous patients) had significantly higher ADP‐induced (78.3 ± 3.4% vs. 57.9 ± 9.3%, p = 0.001), collagen‐induced (78.1 ± 2.9% vs. 56.7 ± 9.3%, p = 0.007), and epinephrine‐induced (76.5 ± 9.2% vs. 59.5 ± 7.70%, p = 0.003) platelet aggregation compared with controls. However, ADP‐, collagen‐, or epinephrine‐induced platelet aggregations were similar to those in obese patients. Platelet aggregation responses to a combination of pretreatment with leptin at concentrations of 20, 50, 100, or 500 ng/mL for 5 minutes and ADP at concentrations of 2 μmol/liter also were evaluated. However, we did not find significant increases in platelet aggregation even at high concentrations of leptin (100 or 500 ng/mL) in leptin‐deficient patients, obese subjects, or controls. Discussion: Our data show that similar to findings in obese humans, homozygous or heterozygous leptin deficiency is associated with increased platelet aggregation compared with controls, and that higher concentrations of leptin do not increase platelet aggregation.     

The effects of biological rhythms and sleep quality on benign paroxysmal positional vertigo and reflux symptom severity

Recently, there has been increased interest in chronotypes and clinical differences between them. However, there is limited information about the potential influence of the chronotypes on clinical manifestations and symptom intensity of somatic diseases. The aim of this study is to evaluate the impact of biological rhythm differences and sleep quality on benign paroxysmal positional vertigo (BPPV) and larengo pharyngeal reflux (LPR) severity. Forty-four LPR patients, 43 BBPV patients and 42 controls were included in the study. The morningness–eveningness questionnaire was used to determine chronotypes, and the Pittsburgh Sleep Quality Index was used to assess subjective sleep quality. Both patient groups reported a significantly greater tendency to eveningness diurnal preferences compared to healthy controls. As with the circadian preferences, patients with BPPV or LPR characterized by poorer sleep quality and worse insomnia than non-patient individuals. It can be concluded that the circadian rhythm and sleep quality are related to the severity of LPR and BPPV.     

Investigation of acetylcholinesterase and mammalian DNA topoisomerases,carbonic anhydrase inhibition profiles,and cytotoxic activity of novel bis(α‐aminoalkyl)phosphinic acid derivatives against human breast cancer

The aim of this study was to evaluate biologically active novel molecules having potentials to be drugs by their antitumor properties and by activities of apoptotic caspase and topoisomerase. Following syntheses of novel eight bis(α‐aminoalkyl)phosphinic acid derivatives ( 4a–h ) as a result of array of reactions, compounds were evaluated by cytotoxic effects in vitro on human breast cancer (MCF‐7) and normal endothelial (HUVEC) cell lines. All phosphinic acid derivatives were effective for cytotoxicity on both MCF‐7 and HUVEC lines, while 4c , 4e , and 4f compounds were found significantly more effective. For the evaluation of antitumor properties of compounds in a highly sensitive method, their effects on inhibiting topoisomerases I and II were investigated. Also, some of the bis(α‐aminoalkyl)phosphinic acid derivatives ( 4a, 4e–h ) showed nice inhibitory action against acetylcholinesterase and human carbonic anhydrase isoforms I and II.     

Factors affecting the adsorption of buchnericin LB, a bacteriocin produced by Lactocobacillus buchneri

Buchnericin-LB adsorbs to gram-positive but not to gram-negative bacteria. The tested gram-positive bacteria were species of Lactobacillus, Pediococcus, Leuconostoc, Enterococcus, Lactococcus, Listeria, Bacillus, Staphylococcus; gram-negative bacteria belonged to the genera Salmonella, Escherichia, Yersinia and Pseudomonas. Buchnericin-LB adsorption depended on pH but not on time and temperature. Also some anions of salts and lipoteichoic acid reduced or inhibited its adsorption. Treatment of cells and cell walls of sensitive bacteria with detergents, organic solvents or enzymes did not affect subsequent binding of buchnericin-LB. Treatment with buchnericin-LB caused sensitive cells to lose high amounts of intracellular K⁺ ions and UV-absorbing materials and became more permeable to o-nitrophenol-β-D-galactopyranoside. Buchnericin-LB (640-2560 AU/ml) decreased the colony forming units (99%) and absorbance values of Listeria monocytogenes and Bacillus cereus. These results indicate that the mode of action of buchnericin-LB is bactericidal and its lethal effect is very rapid.     

Protein kinase D inhibits plasma membrane Na+/H+ exchanger activity

The regulation of plasma membraneNa⁺/H⁺exchanger (NHE) activity by protein kinase D (PKD), a novel proteinkinase C- and phorbol ester-regulated kinase, was investigated. Todetermine the effect of PKD on NHE activity in vivo, intracellular pH(pH_i) measurements were made inCOS-7 cells by microepifluorescence using the pH indicator cSNARF-1.Cells were transfected with empty vector (control), wild-type PKD, orits kinase-deficient mutant PKD-K618M, together with green fluorescentprotein (GFP). NHE activity, as reflected by the rate of acid efflux(J_H), wasdetermined in single GFP-positive cells following intracellularacidification. Overexpression of wild-type PKD had no significanteffect on J_H(3.48 ± 0.25 vs. 3.78 ± 0.24 mM/min in control atpH_i 7.0). In contrast,overexpression of PKD-K618M increasedJ_H (5.31 ± 0.57 mM/min at pH_i 7.0;P < 0.05 vs. control). Transfectionwith these constructs produced similar effects also in A-10 cells,indicating that native PKD may have an inhibitory effect on NHE in bothcell types, which is relieved by a dominant-negative action ofPKD-K618M. Exposure of COS-7 cells to phorbol ester significantlyincreased J_H in control cells but failed to do so in cells overexpressing either wild-type PKD (due to inhibition by the overexpressed PKD) or PKD-K618M(because basal J_Hwas already near maximal). A fusion protein containing the cytosolicregulatory domain (amino acids 637-815) of NHE1 (the ubiquitousNHE isoform) was phosphorylated in vitro by wild-type PKD, but with lowstoichiometry. These data suggest that PKD inhibits NHE activity,probably through an indirect mechanism, and represents a novel pathwayin the regulation of the exchanger.

     

Mechanism of the interaction between ribosomal protein S1 and oligonucleotides.

The interaction of the ribosomal protein S1 from E. coli MRE 600 with oligonucleotides was studied by hydrodynamic, spectrophotometric, and kinetic methods. UV-difference spectra which are induced by the complex formation could be separated into a hyperchromic contribution originating from the nucleic acid moiety and a hypochromic contribution from the protein. Systematic determination of binding and rate constants was carried out by the temperature-jump relaxation technique. From the quantitative evaluation of the relaxation times and the relaxation amplitudes, the following conclusions could be drawn: The stoichiometry of the complex formation is one mole S1 per one mole oligonucleotide. The binding constant K, the recombination rate constant kR, and the dissociation rate constant kD, respectively, were measured at different temperatures. The values at 10 degrees C are K = 2 x 10(6) M-1, kR = 1.3 x 10(8) M-1S-1, kD = 65 s-1 for A(pA) 12 and K = 7.5 x 10(5) M-1, kR = 6.8 x 10(7) M-1S-1, kD = 90 S-1 for U(pU) 12. Discrepancies with data reported elsewhere are discussed. The stacking-unstacking equilibrium of the free oligonucleotide is frozen if the oligonucleotide is bound to the protein. The conformational change of the oligonucleotide does not occur in the form of a preequilibrium, but is induced after the primary binding step.     

AIDS awareness and knowledge among married women living in Malatya (Turkey): implications for province-based prevention programs

The study was performed in 2003 to obtain baseline information on married women's knowledge of AIDS/HIV in Malatya, Turkey. The aim was to reach 1% of the population by covering 1200 married women out of 120,034 whose ages ranged from 15-49. Stratified systematic random sampling was used according to 17 health center lists. Although median HIV/AIDS knowledge score was found to be 70 (highest score was 100), it was also seen that clinical properties of the disease were not recognized adequately. The main resources of knowledge were visual (93%) and printed material (35%). These findings show that media based information does not seem to be sufficient. Therefore, it is recommended that health education should be inserted in the school curriculum and health personnel should inform the women.