Effects of feeding heat-treated colostrum on passive transfer of immune and nutritional parameters in neonatal dairy calves.

The first objective of this study was to describe the effect of on-farm heat treatment of colostrum on colostral bacteria counts and IgG concentrations. The second objective was to describe the effect of feeding heat-treated (vs. raw) colostrum on passive transfer of colostral immune and nutritional parameters in neonatal calves. Pooled batches of colostrum were mixed and divided equally: one half was fed raw whereas the other half was fed after heat treatment at 60 degrees C for 60 min using a commercial on-farm batch pasteurizer. Colostrum samples were cultured for total bacteria count and total coliform count and analyzed for total IgG concentration. Forty-nine Holstein calves were fed either raw colostrum (n = 24) or heat-treated colostrums (n = 25) within 1 to 2 h after birth. Serum samples collected from calves at 0 h (precolostrum) and 24 h (postcolostrum) were assayed for serum total protein; IgG, IgA, and IgM concentrations; peripheral total leukocyte counts; neutrophil counts; lymphocyte counts; lymphocyte phenotypes; vitamin A, vitamin E, cholesterol, and beta-carotene concentrations. Serum samples collected from 2- to 5-d-old calves were tested for immunoglobulin function via a bovine viral diarrhea virus type I serum neutralization titer and for neutrophil bacterial opsonization activity. On-farm batch heat treatment of colostrum at 60 degrees C for 60 min resulted in lower colostrum bacteria concentrations while maintaining colostral IgG concentration. Calves fed heat-treated colostrum had significantly greater serum total protein and IgG concentrations at 24 h, plus greater apparent efficiency of IgG absorption (total protein = 6.3 mg/dL; IgG = 22.3 mg/mL; apparent efficiency of absorption = 35.6%) compared with calves fed raw colostrum (TP = 5.9 mg/dL; IgG = 18.1 mg/mL; apparent efficiency of absorption = 26.1%). There was no effect of treatment on serum concentrations of IgA, IgM, vitamin A, vitamin E, cholesterol, beta-carotene or vitamin E:cholesterol ratio, or on serum bovine viral diarrhea virus type I serum neutralization titers. There was no difference between treatment groups when examining calf plasma total leukocyte counts, neutrophil counts, lymphocyte counts, or neutrophil opsonization activity. However, the latter results were considered inconclusive.

Early and extended early bactericidal activity of levofloxacin, gatifloxacin and moxifloxacin in pulmonary tuberculosis.

OBJECTIVE: To evaluate the early bactericidal activity (EBA) of the new fluoroquinolones levofloxacin, gatifloxacin and moxifloxacin in patients with pulmonary tuberculosis (PTB). DESIGN: Randomized, open-label trial. Forty adults with newly diagnosed smear-positive PTB (10 per arm) were assigned to receive isoniazid (INH) 300 mg, levofloxacin 1000 mg, gatifloxacin 400 mg, or moxifloxacin 400 mg daily for 7 days. Sputum for quantitative culture was collected for 2 days before and daily during 7 days of monotherapy. Bactericidal activity was estimated by measuring the decline in bacilli during the first 2 days (EBA 0-2) and last 5 days of monotherapy (extended EBA, EBA 2-7). Laboratory staff were blinded to treatment assignment. RESULTS: The EBA 0-2 of INH (0.67 log10 cfu/ml/day) was greater than that of moxifloxacin and gatifloxacin (0.33 and 0.35 log10 cfu/ml/day, respectively), but not of levofloxacin 1000 mg daily (0.45 log10 cfu/ml/day) (P = 0.14). Bactericidal activity between days 2 and 7 was similar for all three fluoroquinolones. In a pooled comparison, the EBA 2-7 of the fluoroquinolones was greater than for INH. CONCLUSION: Moxifloxacin, gatifloxacin, and high-dose levofloxacin have excellent EBA, only slightly less than for INH, and greater extended EBA. These drugs warrant further study in the treatment of drug-susceptible TB.

Bioavailability of vitamin D from fortified process cheese and effects on vitamin D status in the elderly.

We conducted 2 studies to determine the effect of vitamin D-fortified cheese on vitamin D status and the bioavailability of vitamin D in cheese. The first study was designed to determine the effect of 2 mo of daily consumption of vitamin D3-fortified (600 IU/d) process cheese on serum 25-hydroxyvitamin D (25-OHD), parathyroid hormone (PTH), and osteocalcin (OC) concentrations among 100 older (> or =60 yr) men and women. Participants were randomized to receive vitamin D-fortified cheese, nonfortified cheese, or no cheese. Serum levels of 25-OHD, PTH, and OC were measured at the beginning and end of the study. There were no differences in 25-OHD, PTH, or OC after 2 mo of fortified cheese intake. The vitamin D-fortified cheese group had a greater decrease in 25-OHD than other groups, due to higher baseline 25-OHD. A second study was conducted to determine whether the bioavailability of vitamin D2 in cheese (delivering 5880 IU of vitamin D2/56.7-g serving) and water (delivering 32,750 IU/250 mL) is similar and whether absorption differs between younger and older adults. The second study was a crossover trial involving 2 groups of 4 participants each (younger and older group) that received single acute feedings of either vitamin D2-fortified cheese or water. Serial blood measurements were taken over 24 h following the acute feeding. Peak serum vitamin D and area under the curve were similar between younger (23 to 50 yr) and older (72 to 84 yr) adults, and vitamin D2 was absorbed more efficiently from cheese than from water. These studies demonstrated that vitamin D in fortified process cheese is bioavailable, and that young and older adults have similar absorption. Among older individuals, consuming 600 IU of vitamin D3 daily from cheese for 2 mo was insufficient to increase serum 25-OHD during limited sunlight exposure.

Molybdopterin synthase mutations in a mild case of molybdenum cofactor deficiency.

Molybdenum cofactor deficiency is a rare inborn error of metabolism with generally severe symptoms, most often including neonatal seizures and severe developmental delay. We describe a patient with an unusually mild form of the disease. Two mutations in MOCS2A (molybdenum cofactor synthesis enzyme 2A) were identified: a single base change, 16C > T, that predicts a Q6X substitution on one allele and a 19G > T transversion that predicts a valine to phenylalanine substitution, V7F, on the second. It is postulated that the milder clinical symptoms result from a low level of residual molybdopterin synthase activity derived from the 19G > T allele.

Duration of efficacy of treatment of latent tuberculosis infection in HIV-infected adults.

BACKGROUND: Treatment of latent infection is needed to protect HIV-infected individuals against tuberculosis. A previous report addressed short-term efficacy of three regimens in HIV-infected adults. We now report on long-term efficacy of the study regimens. METHODS: Three daily self-administered regimens were compared in a randomized placebo-controlled trial in 2736 purified protein derivative (PPD)-positive and anergic HIV-infected adults. PPD-positive subjects were treated with isoniazid (INH) for 6 months (6H), INH plus rifampicin for 3 months (3HR), INH plus rifampicin and pyrazinamide for 3 months (3HRZ), or placebo for 6 months. Anergic subjects were randomized to 6H or placebo. RESULTS: 6H initially protected against tuberculosis in PPD-positive individuals; however, benefit was lost within the first year of treatment. Sustained benefit was observed in persons receiving 3HR and 3HRZ. In a Cox regression analysis, the adjusted relative risk for tuberculosis compared with placebo was 0.67 [95% confidence interval (CI), 0.42-1.07] for 6H, 0.49 (95% CI, 0.29-0.82) for 3HR, and 0.41 (95% CI, 0.22-0.76) for 3HRZ. When the rifampicin-containing regimens were combined, the adjusted relative risk for tuberculosis compared with placebo was 0.46 (95% CI, 0.29-0.71). Among anergic subjects, a modest degree of protection with 6H was present (adjusted relative risk, 0.61; 95% CI, 0.32-1.16). Treatment of latent tuberculosis infection had no effect on mortality. CONCLUSION: Six months of INH provided short-term protection against tuberculosis in PPD-positive HIV-infected adults. Three month regimens including INH plus rifampicin or INH, rifampicin and pyrazinamide provided sustained protection for up to 3 years.

JFC1, a novel tandem C2 domain-containing protein associated with the leukocyte NADPH oxidase.

We have employed a yeast two-hybrid system to screen a B lymphoblast-derived cDNA library, searching for regulatory components of the NADPH oxidase. Using as bait the C-terminal half of p67(phox), which contains both Src homology 3 domains, we have cloned JFC1, a novel human 62-kDa protein. JFC1 possesses two C2 domains in tandem. The C2A domain shows homology with the C2B domain of synaptotagmins. JFC1 mRNA was abundantly expressed in bone marrow and leukocytes. The expression of JFC1 in neutrophils was restricted to the plasma membrane/secretory vesicle fraction. We confirmed JFC1-p67(phox) association by affinity chromatography. JFC1-containing beads pulled down both p67(phox) and p47(phox) subunits from neutrophil cytosol, but when the recombinant proteins were used, only p67(phox) bound to JFC1, indicating that JFC1 binds to the cytosolic complex via p67(phox) without affecting the interaction between p67(phox) and p47(phox). In contrast to synaptotagmins, JFC1 was unable to bind to inositol 1,3,4,5-tetrakisphosphate but did bind to phosphatidylinositol 3,4,5-trisphosphate and to a lesser extent to phosphatidylinositol 3,4-diphosphate. From the data presented here, it is proposed that JFC1 is acting as an adaptor protein between phosphatidylinositol 3-kinase products and the oxidase cytosolic complex.

The desi ways: traditional health practices of South Asian women in Canada.

Although many South Asian immigrants have made their homes in Canada, little research has examined health behaviors in this population and fewer studies have examined the use of traditional health practices. As part of a larger study on health-seeking patterns of South Asian women living in Western Canada, an analysis was done on the use of traditional health practices. Using critical ethnographic methods, data were collected through face-to-face individual interviews (n = 50), focus group discussions (n = 12), and community meetings with a cross section of women in the South Asian community. Interviews were conducted in the language of each participant's choice. Thematic analysis was done on the transcribed interviews. Women's descriptions of traditional health practices varied and consisted of home remedies, dietary regimens, prayers, rituals, and consultation with hakims, veds, babajis, pundits, homeopaths, and jyotshis. Choosing to use traditional health practices was influenced by family members, the nature and severity of problems, beliefs and prior experiences, and the feasibility of using these practices. Traditional health practices were used on a daily or episodic basis. Women rarely used traditional health practices exclusively. Traditional health practices were used for small problems or when conventional medicines did not work. For women to meet their health needs, health care providers must be culturally sensitive and respect women's choices to use traditional health practices.

Hypertonic saline resuscitation abrogates neutrophil priming by mesenteric lymph.

OBJECTIVE: Neutrophil (PMN) priming after hemorrhagic shock is predictive of the subsequent development of multiple organ failure, but the mechanism remains unknown. Recently, we and others have demonstrated that mesenteric lymph from shock animals resuscitated with lactated Ringer's solution (LR) is not only a potent PMN priming agent but also causes lung injury. Work by others has shown that resuscitation with hypertonic saline (HTS) protects animals from lung injury after hemorrhagic shock. Therefore, we hypothesize that resuscitation with HTS will abolish PMN priming by postshock mesenteric lymph. METHODS: After mesenteric lymph duct catheterization, male rats underwent hemorrhagic shock (mean arterial pressure of 40 mm Hg for 90 minutes) and resuscitation with shed blood plus either LR (2x volume of shed blood) or 4 mL/kg of 7% HTS (isonatremic). Priming for superoxide by PMN was measured after fMLP (1 microM) activation. RESULTS: Shock significantly decreased mesenteric lymph flow from preshock levels in both groups. LR resuscitation produced significantly more mesenteric lymph than HTS resuscitation. Mesenteric lymph from LR animals primed PMN for superoxide production, whereas, HTS eliminated this priming. CONCLUSION: HTS not only decreases postshock mesenteric lymph production, it eliminates PMN priming by mesenteric lymph, suggesting a mechanism for the beneficial effects of HTS resuscitation.

Tumour cell kinetics as predictors of response in canine lymphoma treated with chemotherapy alone or combined with whole body hyperthermia.

Kinetic parameters including potential doubling time (Tpot), duration of S phase (Ts), labelling index (LI), and DNA index (DI) were obtained from 42 dogs with previously untreated lymphoma. Standard flow cytometric techniques using BrdUrd were employed. All dogs were treated with L-asparaginase and remission was induced in 26 dogs, which were then randomized to receive chemotherapy only (doxorubicin [DOX] alone or with lonidamine) or chemotherapy plus whole body hyperthermia (WBH). Dogs were treated every 3 weeks for up to five treatments and evaluated every 3 weeks for evidence of tumour recurrence. Within this subset of animals there was no difference in outcome based on treatment group. Median values for Tpot, Ts and LI were 3.4 days, 7.23 h and 12.49%, respectively. Dogs that had tumours with LI > or = 20% had a shorter time until recurrence than dogs with tumours characterized by LI < 20%. In dogs treated only with chemotherapy, dogs bearing tumours with longer than median Tpot and Ts values and lower than median LI had significantly longer remission duration than dogs with more rapidly proliferating tumours. Dogs treated only with chemotherapy, which had longer than median Tpot and Ts values and lower than median LI, had significantly longer remission duration than all other dogs in the study. The mechanisms in which kinetics are associated with response to chemotherapy are not clear and vary depending on tumour type and treatment regimen. More work is needed to understand factors involved in cell killing during in vivo hyperthermia.

Measurement of sputum Mycobacterium tuberculosis messenger RNA as a surrogate for response to chemotherapy.

Effective treatment regimens for pulmonary tuberculosis are difficult to assess because of the slow growth rate of Mycobacterium tuberculosis in culture and its protracted clearance from sputum. A rapid method that reflects effective antimicrobial activity would markedly advance evaluation of treatment and promote the assessment of new antituberculosis drugs. Conventional methods measure the progressive reduction of numbers of acid-fast bacilli in the sputum smear and the clearance of organisms in sputum culture. In this study, we measured levels of M. tuberculosis 85B (alpha antigen) messenger RNA (mRNA), 16S ribosomal RNA (rRNA), and IS6110 DNA in patients' sputa to ascertain whether they could serve as potential surrogate markers of response to chemotherapy. Sputum specimens were sequentially collected for up to a year from 19 smear-positive pulmonary tuberculosis patients receiving an optimal drug treatment regimen. Nucleic acids were isolated from these specimens, and two M. tuberculosis molecular targets (mRNA, DNA) were quantified, using the ABI Prism 7700 Sequence Detection System. The Mycobacterium genus-specific 16S rRNA was quantified with a limiting dilution RT-PCR assay. Results show that levels of 85B mRNA declined after initiation of therapy, as did viable M. tuberculosis colony counts, with 90% of patients becoming negative for both markers after 2 mo of treatment. The rapid disappearance of M. tuberculosis mRNA from sputum suggests that it is a good indicator of microbial viability and a useful marker for rapid assessment of response to chemotherapy.

Dosimetric characteristics of a new high-intensity 192Ir source for remote afterloading.

A new high-intensity 192Ir source has recently become commercially available for remote afterloading brachytherapy treatment. The dosimetric characteristics (dose rate constant, radial dose function, and anisotropy function) of this source were experimentally determined through the application of AAPM Task Group 43 recommendations. Complete dosimetric data are presented in this manuscript.

Isolated sulfite oxidase deficiency.

Isolated sulfite oxidase (SO) deficiency is an autosomal recessively inherited inborn error of sulfur metabolism. In this report of a ninth patient the clinical history, laboratory results, neuropathological findings and a mutation in the sulfite oxidase gene are described. The data from this patient and previously published patients with isolated sulfite oxidase deficiency and molybdenum cofactor deficiency are summarized to characterize this rare disorder. The patient presented neonatally with intractable seizures and did not progress developmentally beyond the neonatal stage. Dislocated lenses were apparent at 2 months. There was increased urine excretion of sulfite and S-sulfocysteine and a decreased concentration of plasma cystine. A lactic acidemia was present for 6 months. Liver sulfite oxidase activity was not detectable but xanthine dehydrogenase activity was normal. The boy died of respiratory failure at 32 months. Neuropathological findings of cortical necrosis and extensive cavitating leukoencephalopathy were reminiscent of those seen in severe perinatal asphyxia suggesting an etiology of energy deficiency. A point mutation that resulted in a truncated protein missing the molybdenum-binding site has been identified.

Phosphorylation of the respiratory burst oxidase subunit p47phox as determined by two-dimensional phosphopeptide mapping. Phosphorylation by protein kinase C, protein kinase A, and a mitogen-activated protein kinase.

The respiratory burst oxidase is responsible for superoxide (O2) production by phagocytes and B lymphocytes. This multicomponent enzyme is dormant in resting cells but is activated on exposure of the cells to an appropriate stimulus. Upon activation, several serine residues on the cytosolic oxidase subunit p47phox become phosphorylated. Using two-dimensional tryptic phosphopeptide mapping, we studied the phosphorylation of p47phox in 32Pi-loaded Epstein-Barr virus-transformed B lymphoblasts expressing wild type p47phox or any of several P47phox Ser -> Ala mutants. We were able to identify the labeled peptides from wild type p47phox as those contain- ing Ser303/304 Ser315, Ser320, Ser328 and/or Ser359/370, and Ser345/348 ; no 32P-labeled Ser310-containing peptide was found. When purified p47phox, was phosphorylated in vitro by various protein kinases, varying phosphopeptide patterns were observed. Protein kinase C phosphorylated all the peptides except the one containing Ser345/348; protein kinase A phosphorylated the peptide containing Ser320 and one or both of the peptides containing Ser328 and Ser359/370; while mitogen-activated protein kinase phophorylated only the peptide containing Ser345/348. These findings suggest that these three kinases play distinct roles in the activation of the respiratory burst oxidase, each of them catalyzing the phosphorylation of a different group of serines in p47phox.

The mechanisms by which hyperbaric oxygen and carbogen improve tumour oxygenation.

Hyperbaric oxygen (HBO) has been proposed to reduce tumour hypoxia by increasing the amount of dissolved oxygen in the plasma. That this actually occurs has not been verified experimentally. This study was performed to explore changes in tumour oxygenation induced by treatment with normobaric and hyperbaric oxygen and carbogen. R3230Ac mammary adenocarcinomas were implanted into Fisher 344 rats. Arterial blood gases, blood pressure and heart rate were monitored. Tumour oxygenation was measured polarographically in five sets of animals. They received either normobaric 100% oxygen, hyperbaric (3 atmospheres; atm) 100% oxygen, normobaric carbogen or hyperbaric (3 atm) carbogen (HBC) +/- bretylium. HBO reduced the mean level of low pO2 values (< 5 mmHg) from 0.49 to 0.07 (P = 0.0003) and increased the average median pO2 from 8 mmHg to 55 mmHg (P = 0.001). HBC reduced the level of low pO2 values from 0.82 to 0.51 (P = 0.002) an increased median pO2 from 2 mmHg to 6 mmHg (P = 0.05). Normobaric oxygen and carbogen did not change tumour oxygenation significantly. Sympathetic blockade with bretylium before HBC exposure improved oxygenation significantly more than HBC alone (low pO2 0.55-0.17, median pO2 4-17 mmHg). HBO and hyperbaric carbogen improved tumour oxygenation in this model, while normobaric oxygen or carbogen had no effect. Sympathetic-mediated vasoconstriction during hyperbaric carbogen caused it to be less effective than HBO. This mechanism also appeared to operate during normobaric carbogen breathing.

Covalently bound phosphate residues in bovine milk xanthine oxidase and in glucose oxidase from Aspergillus niger: a reevaluation.

The reported presence of covalently bound phosphate residues in flavoproteins has significant implications with regard to the catalytic mechanisms and structural stability of the specific enzymes themselves and in terms of general cellular metabolic regulation. These considerations have led to a reevaluation of the presence of covalently bound phosphorus in the flavoproteins xanthine oxidase (xanthine: oxygen oxidoreductase, EC 1.1.3.22) and glucose oxidase (beta-D-glucose: oxygen 1-oxidoreductase, EC 1.1.3.4). Milk xanthine oxidase purified by a procedure that includes anion-exchange chromatography is shown to contain three phosphate residues. All three are noncovalently associated with the protein, two with the FAD cofactor, and one with the molybdenum cofactor. Results of chemical analysis and 31P NMR spectroscopy indicate that enzyme purified by this method contains no phosphoserine residues. Xanthine oxidase preparations purified by chromatography on calcium phosphate gel in place of DEAE-Sephadex yielded higher phosphate-to-protein ratios, which could be reduced to the expected values by additional purification on a folate affinity column. Highly active, highly purified preparations of glucose oxidase are shown to contain only the two phosphate residues of the FAD cofactor. The covalently bound bridging phosphate reported by others may arise in aged or degraded preparations of the enzyme but appears not to be a constituent of functional glucose oxidase. These results suggest that the presence of covalent phosphate residues in other flavoproteins should be rigorously reevaluated as well.

The structure of a molybdopterin precursor. Characterization of a stable, oxidized derivative.

An oxidized pterin species, termed compound Z, has been isolated from molybdenum cofactor-deficient mutants of Escherichia coli and shown to be the direct product of oxidation of a molybdopterin precursor which accumulates in these mutants. The complete structural characterization of compound Z has been accomplished. A carbonyl function at C-1' of the 6-alkyl side chain can be reacted with 2,4-dinitrophenylhydrazine to yield a phenylhydrazone and can be reduced with borohydride, producing a mixture of two enantiomers, each with a hydroxyl group on C-1'. Compound Z contains one phosphate/pterin and no sulfur. The phosphate group is insensitive to alkaline phosphatase and to a number of phosphodiesterases but is quantitatively released as inorganic phosphate by mild acid hydrolysis. From 31P and 1H NMR of compound Z it was inferred that the phosphate is bound to C-2' and C-4' of a 4-carbon side chain, forming a 6-membered cyclic structure. Mass spectral analysis showed an MH+ ion with an exact mass of 344.0401 corresponding to the molecular formula C10H11N5O7P, confirming the proposed structure.

Anthrapyrazole anticancer agents. Synthesis and structure-activity relationships against murine leukemias.

Chromophore modification of the anthracenediones related to mitoxantrone in an attempt to provide agents with diminished or no cardiotoxicity has resulted in a novel class of DNA binders, the anthrapyrazoles. Their synthesis was carried out by a two-stage condensation sequence starting from requisite 1,4- or 1,5-dichloro-9,10-anthracenedione precursors. Reaction with a monoalkylhydrazine gave a chloroanthrapyrazole intermediate whose subsequent condensation with primary or secondary alkylamines provided the target "two-armed" anthrapyrazoles. A-ring 7,10-dihydroxy anthrapyrazoles were derived from amine condensation with intermediate 5-chloro-7,10-dihydroxyanthrapyrazoles or, alternatively, from intermediate 5-chloro-7,10-bis(benzyloxy)anthrapyrazoles followed by hydrogenolysis of the benzyl protecting groups to provide the target compounds. Potent in vitro activity was demonstrated against murine L1210 leukemia in vitro (IC50 = 10(-7)-10(-8) M) as well as against P388 leukemia in vivo over a wide range of structural variants. In general, activity against the P388 line was maximized by basic side chains at N-2 and C-5, two to three carbon spacers between proximal and distal nitrogens of the side chain, and A-ring hydroxylation. Besides having curative activity against the P388 line, the more active compounds were curative against murine B-16 melanoma in vivo. On the basis of their exceptional in vivo anticancer activity, A-ring dihydroxy compounds 71 and 74 reported in this study have been selected for development toward clinical trials.

Synthesis and antileishmanial activity of 6-methoxy-4-methyl-N-[6-(substituted-1-piperazinyl)hexyl]-8-quinolinamines and related compounds.

The 8-quinolinamine, 4-[6-[6-methoxy-4-methyl-8-quinolinyl)amino]hexyl]-1-piperazineethanol (1b), has been shown to be highly effective against Leishmania donovani infections in hamsters. In an effort to obtain a more potent, less toxic 8-quinolinamine, a series of analogues (2) was prepared that examined particularly the structural requirements of the terminal piperazine moiety. Of the substituted piperazines and alternative heterocycles prepared, as well as those quinoline analogues with ring insertion of a methyl group in the 2-position or an aryloxy substituent in the 5-position, an increase in potency was achieved only with the 2-hydroxypropyl analogue (2f).