Erwinia chrysanthemi, a phytopathogenic bacterium, produces a protease inhibitor which is a low-molecular-weight, heat-stable protein. In addition to its action on the three E. chrysanthemi extracellular proteases A, B and C, it also strongly inhibits the 50 kD extracellular protease of Serratia marcescens. Its structural gene (inh) was subcloned and expressed in Escherichia coli, in which it encodes an active inhibitor which was purified. The nucleotide sequence of the inh gene shows an open reading frame of 114 condons. The N-terminal amino acid sequence of the purified inhibitor was also determined. It indicated the existence of an amino-terminal signal peptide absent from the mature protein. The inhibitor is entirely periplasmic in E. chrysanthemi and partially periplasmic in E. coli.

BACKGROUND

In women with polycystic ovary syndrome (PCOS), excess ovarian androgen production is driven by increased LH secretion. Studies conducted in animals suggest that the granulosa cell may influence LH-stimulated theca cell androgen production.

OBJECTIVE

The objective of this study was to determine whether FSH enhances androgen production in women with PCOS compared with that of normal women.

METHODS

A prospective study was conducted to compare androgen production in response to FSH in two groups of women.

METHODS

The study was conducted in a General Clinical Research Center in a tertiary academic medical center.

METHODS

Women with PCOS, 18-35 yr (n = 20), and normal ovulatory controls, 18-35 yr (n = 10), were recruited for study.

METHODS

Serial blood samples were obtained over a 24-h period after an iv injection of recombinant human FSH (150 IU).

METHODS

The main outcome measures were serum 17-hydroxyprogesterone (17-OHP), androstenedione (A), dehydroepiandrosterone (DHEA), testosterone (T), and inhibin B (Inh B) responses after FSH administration.

RESULTS

Basal serum 17-OHP, A, and T levels were markedly increased in women with PCOS compared with that observed in normal women. Basal DHEA and Inh B levels were similar to those of normal controls. After FSH injection, PCOS women demonstrated enhanced production of 17-OHP, A, DHEA, and Inh B, whereas in normal women no increases were observed. T levels declined slightly in both groups.

CONCLUSIONS

These findings provide evidence that, in PCOS women, theca cell androgen production is enhanced by FSH administration and suggest a granulosa-theca cell paracrine mechanism.

We have studied the expression of the complement components C2, C3, factor B, C1 inhibitor (C1-inh), C4-binding protein (C4-bp) and factor H in human peripheral blood monocytes, skin fibroblasts, umbilical vein endothelial cells (HUVEC) and the human hepatoma cell line G2 (Hep G2) in the absence and the presence of interferon-gamma (IFN-gamma). E.l.i.s.a. performed on culture fluids, run-on transcription assays, Northern blot and double-dilution dot-blot techniques confirmed that monocytes expressed all six components, whereas fibroblasts, HUVEC and HepG2 each expressed five of the six components. Fibroblasts and HUVEC did not synthesize C4-bp, and Hep G2 did not produce factor H. In addition to these differences, the synthesis rates of C3, C1-inh and factor H were not the same in all cell types. However, the synthesis rates of C2 and factor B were similar in all four cell types. The half-lives of the mRNAs were shorter in monocytes than in other cell types. Monocyte factor H mRNA had a half-life of 12 min in monocytes, compared with over 3 h in fibroblasts and HUVEC. The instability of factor H mRNA in monocytes may contribute to their low factor H secretion rate. IFN-gamma produced dose-dependent stimulation of C2, factor B, C1-inh, C4-bp and factor H synthesis by all cell types expressing these proteins, but decreased C3 synthesis in all four cell types. Cell-specific differences in the response to IFN-gamma were observed. The increased rates of transcription of the C1-inh and factor H genes in HUVEC were greater than in other cell types, while the increased rate of transcription of the C2, factor B and C1-inh genes in Hep G2 cells was less than in other cell types. IFN-gamma did not affect the stability of C3, factor H or C4 bp mRNAs, but increased the stability of factor B and C1-inh mRNAs and decreased the stability of C2 mRNA. Although these changes occurred in all four cell types studied, the half-life of C1-inh mRNA in monocytes was increased almost 4-fold, whereas the increases in the other cell types were less than 30%. These data show that the constitutive synthesis rates of complement components may vary in the different cell types. They also show that the degree of change in synthesis rates in response to IFN-gamma in each of the cell types often varies due to differences in transcriptional response, sometimes in association with changes in mRNA stability.

The effect of interleukin (IL)-6 and IL-1 on the biosynthesis of complement components C3, factor B, C2, C4 and C1 inhibitor (C1 inh), as well as that of albumin, was studied in vitro in human hepatoma-derived cell line, HepG2. Measuring the amounts of secreted complement proteins we detected a significant upregulation of C3 by both hormones. The enhancement of the factor B and especially that of C1 inh production was predominant by IL-6. In our experimental system neither IL-1 nor IL-6 affected the biosynthesis of C2 and C4. Albumin secretion was significantly decreased only in the simultaneous presence of IL-1 and IL-6. Detection of the changes in the amounts of C3- and factor B-specific mRNA of HepG2 cells suggests a pretranslational regulation by these cytokines. The secretion of C3 and factor B was markedly potentiated when IL-1 and IL-6 were added together. However only the gene expression of factor B, but not of C3, was found to reveal synergism. IL-6 enhanced the in vitro production of C3 in mouse hepatocytes as well. This effect was greatly potentiated in the presence of histamine.

Isoniazid (INH) is one of the most active compounds used to treat tuberculosis (TB) worldwide. In addition, INH has been used as a prophylactic drug for individuals with latent Mycobacterium tuberculosis (MTB) infection to prevent reactivation of disease. Importantly, the definition of multidrug resistance (MDR) in TB is based on the resistance of MTB strains to INH and rifampicin (RIF). Despite its simple chemical structure, the mechanism of action of INH is very complex and involves several different concepts. Many pathways pertaining to macromolecular synthesis are affected, notably mycolic acid synthesis. The pro-drug INH is activated by catalase-peroxidase (KatG), and the active INH products are targeted by enzymes namely, enoyl acyl carrier protein (ACP) reductase (InhA) and beta-ketoacyl ACP synthase (KasA). In contrast, INH is inactivated by arylamine N-acetyltransferases (NATs). Consequently, the molecular mechanisms of INH resistance involve several genes in multiple biosynthetic networks and pathways. Mutation in the katG gene is the major cause for INH resistance, followed by inhA, ahpC, kasA, ndh, iniABC,fadE, furA, Rv1592c and Rv1772. The recent association of efflux genes with INH resistance has also gained considerable attention. Interestingly, substitutions have also been observed in nat, fabD, and accD recently in resistant isolates. Understanding the mechanisms operating behind INH action and resistance would enable better detection of INH resistance. This information would aid novel drug design strategies. Herein we review all mechanisms known to potentially contribute to the complexity of INH action and mechanisms of resistance in MTB, with insights into methods for detection of INH resistance as well as their limitations.

BACKGROUND

Little is known about the incidence of isoniazid-associated hepatitis in HIV-infected Africans who receive both isoniazid preventive therapy (IPT) and antiretroviral therapy (ART).

OBJECTIVE

To assess the rate of and risk factors for isoniazid (INH)-associated hepatitis in persons living with HIV (PLWH) during IPT.

METHODS

PLWH recruited for a clinical trial received 6 months of open-label, daily, self-administered INH at public health clinics. At screening PLWH were excluded if they had any cough, weight loss, night sweats, or other illness. Alcohol abuse was defined as meeting any CAGE criterion. INH-associated hepatitis (INH-hepatitis) was defined as having either alanine or aspartate aminotransferase greater than 5.0 times the upper limit of normal regardless of symptoms when INH was not excluded as the cause.

RESULTS

Of 1,995 PLWH enrolled between 2004 and 2006, 1,762 adhered to at least 4 months of IPT and were analyzed. Nineteen (1.1%) developed hepatitis probably or possibly associated with INH including one death at month 6; 14 of 19 (74%) occurred in months 1-3. Antiretroviral therapy (ART) was received by 480 participants but was not statistically associated with INH-hepatitis (relative risk [RR], 1.56; 95% confidence intervals [CI], 0.62-3.9); those receiving nevirapine had a higher rate (2.0%) than those receiving efavirenz (0.9%; P = 0.34). Although alcohol use did not reach significance (RR, 1.42; 95% CI, 0.57-3.51), meeting at least one CAGE criterion approached statistical significance (RR, 2.37; 95% CI, 0.96-5.84). Neither age greater than 35 years nor the presence of hepatitis B virus core antibody was associated with INH-hepatitis.

CONCLUSIONS

The observed rates of INH-hepatitis were similar to published data. Six months of IPT, which is recommended by the World Health Organization, was relatively safe in this, the largest cohort of African PLWH. Clinical trial registered with www.clinicaltrials.gov (NCT 00164281).

Isoniazid (INH) is an anti-tuberculosis prodrug that is activated by mammalian lactoperoxidase and Mycobacterium tuberculosis catalase peroxidase (MtCP). We report here binding studies, an enzyme assay involving INH, and the crystal structure of the complex of bovine lactoperoxidase (LPO) with INH to illuminate binding properties and INH activation as well as the mode of diffusion and interactions together with a detailed structural and functional comparison with MtCP. The structure determination shows that isoniazid binds to LPO at the substrate binding site on the distal heme side. The substrate binding site is connected to the protein surface through a long hydrophobic channel. The acyl hydrazide moiety of isoniazid interacts with Phe(422) O, Gln(423) O(epsilon1), and Phe(254) O. In this arrangement, pyridinyl nitrogen forms a hydrogen bond with a water molecule, W-1, which in turn forms three hydrogen bonds with Fe(3+), His(109) N(epsilon2), and Gln(105) N(epsilon2). The remaining two sides of isoniazid form hydrophobic interactions with the atoms of heme pyrrole ring A, C(beta) and C(gamma) atoms of Glu(258), and C(gamma) and C(delta) atoms of Arg(255). The binding studies indicate that INH binds to LPO with a value of 0.9 x 10(-6) m for the dissociation constant. The nitro blue tetrazolium reduction assay shows that INH is activated by the reaction of LPO-H(2)O(2) with INH. This suggests that LPO can be used for INH activation. It also indicates that the conversion of INH into isonicotinoyl radical by LPO may be the cause of INH toxicity.

Toll-like receptor (TLR)9 performs our innate response to bacterial DNA, warning us of the presence of infection. Inhibitory oligodeoxyribonucleotides (INH-ODN) have been developed that selectively block activation of mouse TLR9. Their inhibitory motif consisting of CCx(not-C)(not-C)xxGGG (x = any base) also reduces anti-DNA antibodies in lupus mice. The current study demonstrates that this motif also provides the sequences required to block TLR9 in human B cells and human embryonic kidney (HEK) cells transfected with human TLR9. However, extending the sequence by four to five bases at the 5' end enhanced activity and this enhancement was greater when a phosphorothioate (pS) backbone replaced the native phosphodiester (pO) backbone. A series of pO-backbone INH-ODN representing a 500-fold range of activity in biologic assays was shown to cover less than a 2.5-fold range of avidity for binding human TLR9-Ig fusion protein, eliminating TLR9 ectodomain binding as the explanation for sequence-specific differences in biologic activity. With few exceptions, the relative activity of INH-ODN in Namalwa cells and HEK/human TLR9 cells was similar to that seen in mouse B cells. INH-ODN activity in human peripheral blood B cells correlated significantly with the cell line data. These results favor the conclusion that although the backbone determines strength of TLR9 binding, critical recognition of the INH-ODN sequence necessary for biologic activity is performed by a molecule that is not TLR9. These studies also identify the strongest INH-ODN for human B cells, helping to guide the selection of INH-ODN sequences for therapeutics in any situation where inflammation is enhanced by TLR9.

Conventional indirect drug susceptibility testing of Mycobacterium tuberculosis with liquid medium is well established and offers time-saving and reliable results. This multicenter study was carried out to evaluate if drug susceptibility testing (DST) can be successfully carried out directly from processed smear-positive specimens (direct DST) and if this approach could offer substantial time savings. Sputum specimens were digested, decontaminated, and concentrated by the laboratory routine procedure and were inoculated in Bactec MGIT 960 as well as Lowenstein-Jensen (LJ) medium for primary isolation. All the processed specimens which were acid-fast bacterium (AFB) smear positive were used for setting up direct DST for isoniazid (INH) and rifampin (RIF). After the antimicrobial mixture of polymyxin B, amphotericin B, nalidixic acid, trimethoprim, and azlocillin (PANTA) was added, the tubes were entered in the MGIT 960 instrument using the 21-day protocol (Bactec 960 pyrazinamide [PZA] protocol). Results obtained by direct DST were compared with those obtained by indirect DST to establish accuracy and time savings by this approach. Of a total of 360 AFB smear-positive sputum specimens set up for direct DST at four sites in three different countries, 307 (85%) specimens yielded reportable results. Average reporting time for direct DST was 11 days (range, 10 to 12 days). The average time savings by direct DST compared to indirect DST, which included time to isolate a culture and perform DST, was 8 days (range, 6 to 9 days). When results of direct DST were compared with those of indirect DST, there was 95.1% concordance with INH and 96.1% with rifampin. These findings indicate that direct DST with the Bactec MGIT 960 system offers further time savings and is a quick method to reliably detect multidrug resistance (MDR) cases.

BACKGROUND

Few studies have examined predictors of latent tuberculosis infection (LTBI) treatment completion in inner city populations in the United States.

OBJECTIVE

To assess LTBI treatment completion rates and predictors in an inner city cohort.

METHODS

Data from control groups of two sequentially conducted randomized controlled trials of LTBI treatment were analyzed for treatment completion rates. Participants in Study A (n = 191), conducted in 1996-1999, self administered daily isoniazid (INH) for 6-12 months, while participants in Study B (n = 123), conducted in 2002-2005, self administered daily INH for 9 months.

RESULTS

Overall, 44.6% of participants completed therapy, with significantly higher completion rates in Study B than Study A (37.0% vs. 56.1%, P = 0.001). Marriage and alcohol use were significant predictors of completion (aOR = 2.153, 95%CI 1.301-3.562) and non-completion (aOR = 0.530, 95%CI 0.320-0.877), respectively; multivariate analysis indicated increased completion among married persons of foreign birth and among alcohol users who were homeless. Knowledge of and attitudes to tuberculosis were not significant predictors.

CONCLUSIONS

The design provided an opportunity to assess predictors of LTBI treatment completion in this inner city population. Social circumstances were the strongest predictors of treatment completion, suggesting that tangible social services may be more effective than educational programs in encouraging treatment completion.

Previously, we presented in vivo evidence for a physiological significance of cAMP-regulated CFTR Cl(-) channels in Ca(2+)-activated Cl(-) reabsorption in the ductal system of the rat submandibular gland. Here, we address the mechanism by which basal CFTR activation contributes to the transepithelial Cl(-) movement evoked by muscarinic stimulation. The Cl(-) concentration ([Cl(-)]) was increased in the final saliva from rat submandibular gland during pilocarpine stimulation when a small interfering RNA for CFTR or a specific CFTR inhibitor, CFTR(inh)-172, was injected retrogradely into the gland's own duct, indicating that basal CFTR activation is involved in Cl(-) reabsorption. Systemically administered propranolol failed to alter the [Cl(-)], suggesting little involvement of a beta-adrenergic pathway in the Cl(-) movement that occurs through basal CFTR activation. Intraductal injection of suramin (a nonspecific P2-receptor antagonist) increased the salivary [Cl(-)], indicating the existence of endogenous purinergic activation. Upon separate intraductal injection, ATP and a P2Y(2)-receptor agonist, UTP, decreased the salivary [Cl(-)] almost equipotently. CFTR(inh)-172 and suramin each prevented these effects, whereas 2',3'-O-(4-benzoylbenzoyl)-ATP (Bz-ATP), a P2X(7) agonist, had no specific effect. Pilocarpine stimulation evoked ATP secretion into the salivary fluid. Immunohistochemistry revealed the partial coexistence of CFTR and P2Y(2) receptors on the luminal surface of epithelial cells in the striated ducts of this gland. These results raise the possibility that muscarinic stimulation-induced Cl(-) reabsorption occurs through basal CFTR activity and that this is regulated by P2Y(2) receptors in the ductal epithelium via stimulation by ATP secreted into the salivary fluid.

The crystal structure of the complex between the 50 kDa metallo-endoproteinase from Serratia marcescens (SMP), a member of the metzincin superfamily, and an inhibitor from Erwinia chrysanthemi (Inh) was solved by molecular replacement using the known structure of SMP, and refined at 2.30 A resolution to a crystallographic R-factor of 0.195. The E. chrysanthemi inhibitor folds into a compact eight-stranded antiparallel beta-barrel of simple up-down topology such as is found for members of the retinol binding protein family. It mainly interacts with the protease via its five N-terminal residues, which insert into the active site cleft, occupying the S' sites. The first N-terminal residue, SerI1, is partially cleaved off by the protease, while SerI2 makes a hydrogen bond with the catalytically active glutamic acid, Glu177, of the protease. Further interactions are made between one face of the inhibitor formed by the strands s3, s4 and s5 and the protease segment 218 to 228, which is located immediately after the characteristic "Met-turn" of the metzincins.

BACKGROUND

Liver diseases are common in patients with HIV due to viral hepatitis B and C co-infections, opportunistic infections or malignancies, antiretroviral drugs and drugs for opportunistic infections.

OBJECTIVE

To describe the spectrum of liver diseases in HIV-infected patients attending an HIV clinic in Kampala, Uganda.

METHODS

Consecutive patients presenting with jaundice, right upper quadrant pain with fever or malaise, ascites and/or tender hepatomegaly were recruited and underwent investigations to evaluate the cause of their liver disease.

RESULTS

Seventy-seven consecutive patients were recruited over an eleven month period. Of these, 23 (30%) had increased transaminases because of nevirapine (NVP) and/or isoniazid (INH) hepatotoxicity. Although 14 (61%) patients with drug-induced liver disease presented with jaundice, all recovered with drug discontinuation. Hepatitis B surface antigen was positive in 11 (15%) patients while anti-hepatitis C antibody was reactive in only 2 (3%). Probable granulomatous hepatitis due to tuberculosis was diagnosed in 7 (9%) patients and all responded to anti-TB therapy. Other diagnoses included alcoholic liver disease, AIDS cholangiopathy, hepatocellular carcinoma, schistosomiasis, haemangioma and hepatic adenoma. Twelve (16%) patients died during follow-up of which 7 (9%) died because of liver disease.

CONCLUSIONS

Drug history, liver enzyme studies, ultrasound, and hepatitis B and C investigations identified the probable etiology in 60 (78%) of 77 patients with HIV infection presenting with symptoms and/or signs of liver disease.

It is well established in many mammalian species, including the horse that normal testicular function is dependent upon a functional hypothalamic-pituitary-testicular (HPT) axis, which involves classic feedback mechanisms. The major HPT hormones involved in the stallion are gonadotropin-releasing hormone (GnRH), luteinizing hormone (LH), follicle stimulating hormone (FSH), testosterone (T), estrogens (Es) and inhibin (INH). Although prolactin (PRL) fluctuates with season in the stallion and both PRL and thyroid hormone (TH) affect reproduction in other male species, their effects on stallion reproduction have not been elucidated. Growth hormone (GH) in the stallion may be involved in sperm motility, production and secretion of insulin-like growth factor-1 (IGF-1) and LH-induced testosterone release. The action of these hormones and the products involved for normal spermatogenesis require cell to cell communication within the testis. The somatic cell types, Leydig, Sertoli and peritubular myoid cells, all support germ cell development, maturation and release into the seminiferous tubule lumen. The cell to cell crosstalk involves an intricate network of paracrine-autocrine systems that support the endocrine input to modulate cell function. In other male species, researchers have demonstrated the reproductive effects of such paracrine-autocrine factors as IGF-1, transferrin, androgens, estrogens, inhibin, insulin like peptide 3 (INSL3), beta-endorphin and oxytocin. The specific nature and relative contribution of these various factors on testicular function in fertile and subfertile stallions are under investigation. This review summarizes current information regarding the nature of the multiple endocrine-paracrine-autocrine systems that may be necessary for normal testicular function in the stallion.

BACKGROUND

Serum FSH levels rise with increasing age in normal women, particularly as they enter the menopausal transition and progress to the postmenopausal state. The contributions of decreasing levels of inhibin-A (INH-A) and inhibin-B (INH-B) to this rise are presently unclear, as there are no reports of dimeric INH levels in relation to menopausal status. The present study was undertaken in order to provide preliminary data on relationships amongst the dimeric inhibins, oestradiol (E2) and FSH in normal subjects of defined menopausal status.

METHODS

Single serum samples were obtained between cycle days 3 and 8 in regularly cycling women, or at random in those with irregular cycles or amenorrhoea, in 110 women, aged 48-59 years, in the third year of a prospective longitudinal study of the menopausal transition, 'The Melbourne Women's Mid-Life Health Project'. Samples were assayed for FSH, E2, INH-A, INH-B and immunoreactive inhibin (IR-INH) and results were analysed following logarithmic transformation. Undetectable values were assigned the limit of sensitivity of the respective assays. The relationships between hormones were evaluated as a function of menopausal stage. The latter was assigned as Stage 1, premenopausal (no reported change in menstrual cycle pattern), Stage 2, early peri-menopausal (reported change in menstrual cycle frequency in the preceding year with a bleed in the preceding 3 months), Stage 3, late peri-menopausal (no menses in the preceding 3-11 months) and Stage 4, postmenopausal (no menses in the preceding 12 months).

RESULTS

The hormone concentrations in premenopausal subjects (geometric means, FSH 13.5 IU/l, E2 306 pmol/l, IR-INH 217 U/l, INH-A 96 ng/l, and INH-B 48 ng/l) were used as reference points for the other stages of menopausal status. Early peri-menopausal subjects had significantly lower levels of IR-INH (147 U/l) and INH-B (13.5 ng/l) in the presence of a small, statistically nonsignificant rise in FSH (to 21.4 U/l) and no significant change in E2 or INH-A. In late peri-menopausal subjects, IR-INH fell to 76 U/l, INH-A fell to 4.2 ng/l, whilst INH-B was not significantly different at 14 ng/l. FSH had risen significantly to 72.21 U/l. Oestradiol also fell significantly to 89 pmol/l. In the postmenopausal subjects there were no further significant changes in the peptide hormones or FSH, but E2 fell further to 41 pmol/l. There was a significant (P < 0.05) inverse correlation between FSH and E2 (R = -0.78), FSH and IR-INH (R = -0.66), FSH and INH-A (R = -0.53), FSH and INH-B (R = -0.29) while IR-INH and either INH-A or INH-B were positively correlated (R = +0.57 and +0.35, respectively). The data are consistent with negative feedback roles for both dimeric inhibins and E2 as contributors to the regulation of FSH secretion as menopausal status changes.

CONCLUSIONS

The major significant endocrine event in women in the early peri-menopausal phase of the menopausal transition is a substantial fall in the circulating levels of inhibin-B with no significant change in inhibin-A or oestradiol. Progression to late peri-menopausal status is accompanied by a marked fall in inhibin-A and oestradiol and a rise in FSH without further change in inhibin-B. Inhibin-B, a marker of follicle number, is a significant factor in the endocrinology of the menopausal transition.

METHODS

Low serum concentrations of anti-tuberculosis drugs have occasionally been associated with treatment failure.

OBJECTIVE

To determine the prevalence of low serum concentrations of anti-tuberculosis drugs and to identify the determinants of drug concentrations.

METHODS

Venous blood was obtained 2 h after drug ingestion, and serum levels of isoniazid (INH), rifampicin (RMP), ethambutol (EMB), pyrazinamide (PZA), acetyl INH and 25-desacetyl RMP were analysed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Patients with human immunodeficiency virus co-infection and gastrointestinal disease or diarrhoea were excluded.

RESULTS

Among 69 enrolled TB patients, the prevalence of a low 2 h serum concentration of at least one anti-tuberculosis drug was 46.4%. Prevalences of a low concentration of INH, RMP, EMB or PZA were 15.2%, 23.5%, 22.4% and 4.5%, respectively. By multivariate linear regression analysis, the serum concentrations of INH, RMP and PZA were positively associated with dose per kg of body weight (P < 0.05). Moreover, INH concentration was associated with acetyl INH/INH ratio (beta = -8.588, P < 0.001) and EMB concentration was associated with calculated creatinine clearance (beta = -0.025, P < 0.001).

CONCLUSIONS

Low concentrations of anti-tuberculosis drugs are common, and although the clinical significance of low concentrations remains uncertain, it may be necessary to optimise drug doses by therapeutic drug monitoring, especially in patients with an inadequate clinical response to chemotherapy.

Murine zymosan-induced peritonitis represents a well-defined model of acute inflammation. However, the molecular mechanisms by which leukocytes degrade basement membranes during extravasation into the peritoneum are not clear. Gelatinase B (MMP-9) is thought to participate in cellular migration, yet its role in leukocyte transmigration through endothelia during inflammation remains controversial. The aim of the present study was to evaluate the role of MMP-9 in the cell influx during zymosan-induced experimental peritonitis. In zymosan-treated Balb/c mice MMP-9 and its natural inhibitor (tissue inhibitor of metalloproteinase 1 - TIMP-1) were present in the peritoneal fluid and plasma at the time of peritoneal neutrophil (polymorphonuclear leukocyte - PMN) infiltration and persisted there until the time of monocytes/macrophages influx. To probe the function of gelatinases, gelatinase B-deficient mice (MMP-9(-/-)) were used as well as Balb/c mice treated with cyclic CTTHWGFTLC (INH), a specific peptide inhibitor of gelatinases. The studies revealed that in either group of mice deprived of MMP-9 activity, PMN infiltration was impaired at the time of their maximal extravasation (6h) while tumor necrosis factor alpha (TNF-alpha), cytokine-induced neutrophil chemoattractant (KC) and interleukin 10 (IL-10) levels were not changed. At later stages (24 h post-zymosan) a significant increase in PMNs was observed in MMP-9(-/-) mice, but not in the inhibitor-treated mice, in comparison to their respective controls. Moreover, intraperitoneal (i.p.) injection of recombinant mouse pro-MMP-9 induced leukocyte accumulation in peritoneum. Collectively, the findings indicate that gelatinase B participates in leukocyte transmigration; however, its function can be compensated by other mechanisms.

Activation of poly(ADP-ribose) synthetase (PARS, also termed polyADP-ribose polymerase or PARP) has been proposed as a major mechanism contributing to beta-cell destruction in type I diabetes. In the present study, we have investigated the role of PARS in mediating the induction of diabetes and beta-cell death in the multiple-low-dose-streptozotocin (MLDS) model of type I diabetes. Mice genetically deficient in PARS were found to be less sensitive to MLDS than wild type mice, with a lower incidence of diabetes and reduced hyperglycemia. A potent inhibitor of PARS, 5-iodo-6-amino-1,2-benzopyrone (INH(2)BP), was also found to protect mice from MLDS and prevent beta-cell loss, in a dose-dependent manner. Paradoxically, in the PARS deficient mice, the compound increased the onset of diabetes. In vitro the cytokine combination; interleukin-1beta, tumor necrosis factor-alpha and interferon-gamma inhibited glucose-stimulated insulin secretion from isolated rat islets of Langerhans and decreased RIN-5F cell viability. The PARS inhibitor, INH(2)BP, protected both the rat islets and the beta-cell line, RIN-5F, from these cytokine-mediated effects. These protective effects were not mediated by inhibition of cytokine-induced nitric oxide formation. Inhibition of PARS by INH(2)BP was unable to protect rat islet cells from cytokine-mediated apoptosis. Cytokines, peroxynitrite and streptozotocin were all shown to induce PARS activation in RIN-5F cells, an effect suppressed by INH(2)BP. The present study provides evidence for in vivo PARS activation contributing to beta-cell damage and death in the MLDS model of diabetes, and indicates a role for PARS activation in cytokine-mediated depression of insulin secretion and cell viability in vitro.

BACKGROUND

Percent total body surface area (TBSA) burn, inhalation injury (INH), and age all have been shown to be independent predictors of mortality in burn victims. Little is known regarding patients sustaining combined thermal and mechanical injuries in relation to either injury sustained in isolation or with regard to these variables. This descriptive study profiles the 10-year experience of a single American Burn Association/American College of Surgeons verified Level I trauma and burn center and the treatment of this patient population.

METHODS

A retrospective review of all burn and trauma patients admitted between 1990 and 2000. Patients were divided into three groups; Burn only (B), Trauma only (T), and combined Burn/Trauma (B/T). Groups were compared with respect to age, TBSA burn, length of stay (LOS), Injury Severity Score (ISS), INH and mortality. These groups were then compared with B, T and B/T patients from the National Burn Repository (NBR) and National Trauma Data Bank (NTDB). Student's t test and chi tests were performed, as well as multiple logistic regression to identify independent predictors of mortality. p <0.05 was considered significant.

RESULTS

Through our trauma registry, 24,093 patients were identified (T=22,284, B=1717 and B/T=92). When comparing B and T, there was no difference in age, LOS, ISS, or mortality to those patients in the NBR or NTDB. B/T patients showed significantly increased percentage with INH (B/T=44.5% versus 11%), increased LOS (B/T=18 days versus 13.7 B and 5.3 T) and increased mortality (B/T=28.3% versus 9.8% B and 4.3% T). B/T were also significantly older (B/T=40.1 years versus 31.0 B and 35.1 T). When these variables are compared with the NBR and the NTDB benchmarks, mortality (28.3% versus 11.6% NBR and 7.0% NTDB) and ISS (23 versus 11.7 NTDB) were significantly higher with no difference in age (40.1 versus 33.4 NTDB, 35.9 NBR), LOS (18 days versus 23.3 NBR) or TBSA (20.8% versus 19.5% NBR). Multiple logistic regression comparing TBSA, age, ISS and INH of survivors versus non-survivors identified only ISS as an independent predictor of mortality.

CONCLUSIONS

B combined with T presents a rare injury pattern that has a synergistic effect on mortality. Physicians and caregivers should be aware of a 2-3 fold increase in the incidence of INH in this population, and increased mortality despite similar TBSA burned when compared with patients with B as the sole mechanism; ISS appears to be an independent predictor of mortality in this combined injury pattern.

OBJECTIVE

We sought to determine the association of abnormal second-trimester serum analytes with early preterm preeclampsia.

METHODS

We conducted a retrospective study of 7767 subjects undergoing second-trimester serum aneuploidy screening. Values of maternal serum α-fetoprotein (AFP), β-human chorionic gonadotropin (hCG), and inhibin (INH) were calculated as multiples of the median (MoM) and evaluated by gestational age at delivery and occurrence of preeclampsia.

RESULTS

Of 459 (6.5%) cases of preeclampsia, 65 (14%) delivered <34 weeks and 394 (86%) delivered >34 weeks. Elevated AFP, hCG, and INH >2 MoM were associated with preeclampsia, and the odds ratio was higher for the development of preeclampsia <34 weeks than >34 weeks (odds ratio, 8.04 vs 2.91 for AFP, 3.6 vs 2 for hCG, and 4.17 vs 3.08 for INH, P < .001 for all). The higher the MoM for each analyte the greater the likelihood of preeclampsia.

CONCLUSIONS

Elevated AFP, hCG, and INH levels >2 MoM are associated with developing early preeclampsia, and the more elevated they are, the higher the likelihood.

BACKGROUND

Infectious diseases remain among the major morbid events in patients with end-stage renal disease (ESRD) on renal replacement therapy (RRT). In developing countries, tuberculosis (TB) has been found to occur more frequently in these patients than in the general population. Efficacy of isoniazid (INH) chemoprophylaxis has been seen in other situations, such as human immunodeficiency virus infection. However, studies on INH prophylaxis in ESRD patients on RRT are limited.

METHODS

In this prospective randomized controlled trial, from April 2000 to June 2001, a total of 109 ESRD patients registered for renal transplant and accepted for maintenance hemodialysis in our hospital were included and followed up until June 2004 to assess the role of INH prophylaxis in preventing development of TB. At the time of acceptance for hemodialysis, 54 patients were assigned to receive daily INH for 1 year and 55 patients were assigned to the control group. Primary outcome was development of TB. Secondary outcome was INH hepatotoxicity. To evaluate the effect of INH prophylaxis on the development of TB, a Kaplan-Meier survival estimate was used to plot TB-free survival curve and log-rank test was used for comparison.

RESULTS

Overall, TB was diagnosed in 27 patients during RRT, with an incidence of 24.8%. TB developed in 9 (16.7%) patients in the INH group and in 18 (32.7%) patients in the control group. There was a significantly lower incidence of TB in the INH group as compared with the control group. The risk ratio of INH vs. control group for development of TB was 0.40 (95% confidence index [CI], 0.17-0.92; P=0.032). In the INH group 27 (50%) patients and in the control group 17 (30.9%) patients developed some hepatic dysfunction. However, significant hepatitis that required discontinuation of INH developed in only 9 (16.7%) patients in the INH group. Furthermore, significant hepatitis also developed in 6 (10.9%) patients in the control group. The majority of patients with significant hepatitis in both groups (INH as well as control) were subsequently found to be positive for hepatitis B and/or hepatitis C viral infection. Mild hepatitis (which did not require discontinuation of INH) was seen in 18 (33.3%) patients in the INH group and 11 (20%) patients in the control group. Viral hepatitis infection was not found in any of the milder cases of hepatitis in either group.

CONCLUSIONS

This study shows significant efficacy of INH chemoprophylaxis during RRT in preventing development of TB, when the INH was started during dialysis itself. INH chemoprophylaxis was safe and well tolerated in the majority of patients. However, mild hepatic dysfunction was common, both in the treatment as well as in the control group. As the incidence of viral hepatitis overall was high in our patients on RRT, it is difficult to identify INH-induced hepatitis in this clinical setting.

Activin A is a member of the transforming growth factor beta (TGF-beta) superfamily and is a strong differentiation factor of embryonic stem (ES) cells. It is unknown whether activin A has any correlation with carcinoma cell differentiation. We investigated the expression of activin-betaA (Act-betaA) which is a subunit of activin A, its receptor type I and IIb (ActRI, ActRIIb) and its inhibitor, inhibin-alpha (Inh-alpha), which is a subunit of inhibin A in esophageal carcinoma by reverse transcription polymerase chain reaction (RT-PCR) method. Act-betaA was overexpressed in carcinoma tissues significantly (p=0.030). On the other hand, Inh-alpha, ActRI and ActRIIb were neither overexpressed, nor suppressed. In immunohistochemistry and in situ hybridization analysis, Act-betaA expression was mainly derived from carcinoma cells. The mRNA expression of Act-betaA was not associated with carcinoma cell differentiation but lymph node metastasis (n0, 1, 2 vs. n3, 4; p=0.013) and clinical stage (I, II, III vs. IV; p=0.026). Moreover, patients with high mRNA expression of Act-betaA had a tendency to show poor prognosis compared to those with low mRNA expression (p=0.064). The finding indicated that activin A expression might not be associated with carcinoma cell differentiation but tumor aggressiveness such as lymph node metastasis in esophageal carcinoma.

BACKGROUND

Immigrants to the U.S. are required to undergo overseas screening for tuberculosis (TB), but the value of evaluation and treatment following entry to the U.S. is not well understood. We determined the cost-effectiveness of domestic follow-up of immigrants identified as tuberculosis suspects through overseas screening.

METHODS

Using a stochastic simulation for tuberculosis reactivation, transmission, and follow-up for a hypothetical cohort of 1000 individuals, we calculated the incremental cost-effectiveness of follow-up and evaluation interventions. We utilized published literature, California Reports of Verified Cases of Tuberculosis (RVCTs), demographic estimates from the California Department of Finance, Medicare reimbursement, and Medi-Cal reimbursement rates. Our target population was legal immigrants to the United States, our time horizon is twenty years, and our perspective was that of all domestic health-care payers. We examined the intervention to offer latent tuberculosis therapy to infected individuals, to increase the yield of domestic evaluation, and to increase the starting and completion rates of LTBI therapy with INH (isoniazid). Our outcome measures were the number of cases averted, the number of deaths averted, the incremental dollar cost (year 2004), and the number of quality-adjusted life-years saved.

RESULTS

Domestic follow-up of B-notification patients, including LTBI treatment for latently infected individuals, is highly cost-effective, and at times, cost-saving. B-notification follow-up in California would reduce the number of new tuberculosis cases by about 6-26 per year (out of a total of approximately 3000). Sensitivity analysis revealed that domestic follow-up remains cost-effective when the hepatitis rates due to INH therapy are over fifteen times our best estimates, when at least 0.4 percent of patients have active disease and when hospitalization of cases detected through domestic follow-up is no less likely than hospitalization of passively detected cases.

CONCLUSIONS

While the current immigration screening program is unlikely to result in a large change in case rates, domestic follow-up of B-notification patients, including LTBI treatment, is highly cost-effective. If as many as three percent of screened individuals have active TB, and early detection reduces the rate of hospitalization, net savings may be expected.

BACKGROUND

CMI is caused by chronic occlusive disease of mesenteric arteries. In such an uncommon disease, clear recommendations are strongly needed. Unfortunately, treatment options for symptomatic CMI are still controversial and no guidelines exist.

METHODS

A systematic literature review of the last 25-years was conducted through MEDLINE, Embase, and Cochrane Review/Trials register to identify studies reporting on CMI treatment with more than 10 patients. Primary outcomes were perioperative mortality and morbidity rates. Secondary outcomes were survival rates, primary and secondary patency rates, vessels treated, CMI recurrence, follow-up (FU), technical success (TS), and in-hospital length of stay (InH-LOS). Patients were divided into endovascular treatment (ET) or open treatment (OT) groups. Subsequently, primary and secondary outcomes were analyzed by study publication year for the interval periods 1986-2000 ("A") and 2001-2010 ("B"). Differences were assessed using the t-test and the χ(2) test.

RESULTS

Forty-three articles with 1,795 patients were included. Perioperative mortality and morbidity rates were lower in the ET group. No difference in survival rate was observed. Primary and secondary patencies were superior in the OT group. A greater number of vessels were revascularized in the OT group. CMI recurrence was more frequent in the ET group. FU was longer in the OT group. TS was superior in the OT group and InH-LOS was shorter in the ET group. A higher number of patients were treated by ET in the period "A." No differences in mortality and morbidity were observed between period "A" and "B" in ET and OT groups.

CONCLUSIONS

Considering the lower periprocedural mortality and morbidity after ET, this approach should be considered as the first treatment option in most CMI patients, especially in those with severe malnutrition. Primary OT should be restricted to cases that do not qualify for ET or good surgical risk patients with long life expectancy. Considering better long-term results of OT, ET treatment should be considered as a bridge therapy to OT in some patients requiring retreatment if ET does not preclude subsequent OT.