Biomedical subjects

R Meyer

Publications and source records attributed to R Meyer.

At least 19 recordsLinked to original sources

PubMed2006

Construction and partial characterization of a Corynebacterium pseudotuberculosis bacterial artificial chromosome library through genomic survey sequencing.

Corynebacterium pseudotuberculosis is a gram-positive bacterium that causes caseous lymphadenitis in sheep and goats. However, despite the economic losses caused by caseous lymphadenitis, there is little information about the molecular mechanisms of pathogenesis of this bacterium. Genomic libraries constructed in bacterial artificial chromosome (BAC) vectors have become the method of choice for clone development in high-throughput genomic-sequencing projects. Large-insert DNA libraries are useful for isolation and characterization of important genomic regions and genes. In order to identify targets that might be useful for genome sequencing, we constructed a C. pseudotuberculosis BAC library in the vector pBeloBAC11. This library contains about 18,000 BAC clones, with inserts ranging in size from 25 to 120 kb, theoretically representing a 390-fold coverage of the C. pseudotuberculosis genome (estimated to be 2.5-3.1 Mb). Many genomic survey sequences (GSSs) with homology to C. diphtheriae, C. glutamicum, C. efficiens, and C. jeikeium proteins were observed within a sample of 215 sequenced clones, confirming their close phylogenetic relationship. Computer analyses of GSSs did not detect chimeric, deleted, or rearranged BAC clones, showing that this library has low redundancy. This GSSs collection is now available for further genetic and physical analysis of the C. pseudotuberculosis genome. The GSS strategy that we used to develop our library proved to be efficient for the identification of genes and will be an important tool for mapping, assembly, comparative, and functional genomic studies in a C. pseudotuberculosis genome sequencing project that will begin this year.

Chromosomes, Artificial, Bacterial↗

PubMed2006

Second malignancy risk associated with treatment of Hodgkin's lymphoma: meta-analysis of the randomised trials.

BACKGROUND: Despite several investigations, second malignancy risks (SMR) following radiotherapy alone (RT), chemotherapy alone (CT) and combined chemoradiotherapy (CRT) for Hodgkin's lymphoma (HL) remain controversial. PATIENTS AND METHODS: We sought individual patient data from randomised trials comparing RT versus CRT, CT versus CRT, RT versus CT or involved-field (IF) versus extended-field (EF) RT for untreated HL. Overall SMR (including effects of salvage treatment) were compared using Peto's method. RESULTS: Data for between 53% and 69% of patients were obtained for the four comparisons. (i) RT versus CRT (15 trials, 3343 patients): SMR were lower with CRT than with RT as initial treatment (odds ratio (OR) = 0.78, 95% confidence interval (CI) = 0.62-0.98 and P = 0.03). (ii) CT versus CRT (16 trials, 2861 patients): SMR were marginally higher with CRT than with CT as initial treatment (OR = 1.38, CI 1.00-1.89 and P = 0.05). (iii) IF-RT versus EF-RT (19 trials, 3221 patients): no significant difference in SMR (P = 0.28) although more breast cancers occurred with EF-RT (P = 0.04 and OR = 3.25). CONCLUSIONS: Administration of CT in addition to RT as initial therapy for HL decreases overall SMR by reducing relapse and need for salvage therapy. Administration of RT additional to CT marginally increases overall SMR in advanced stages. Breast cancer risk (but not SMR in general) was substantially higher after EF-RT. Caution is needed in applying these findings to current therapies.

Combined Modality Therapy↗

PubMed2006

Role of Toll-like receptor 4 for the pathogenesis of acute lung injury in Gram-negative sepsis.

BACKGROUND AND OBJECTIVE: Proinflammatory cytokines as well as nitric oxide (NO) play a major role in mediating the response to lipopolysaccharide (LPS). The present study tested the hypothesis that LPS induces proinflammatory cytokines in the lung via the Toll-like receptor 4 (TLR4)/CD14 signalling cascade. METHODS: Control mice and TLR4-deficient (TLR4-D) mice were used to test TLR4-mediated effects of LPS. Both strains received either Escherichia coli LPS (20 mg kg-1 intraperitoneal) or saline and their lungs were collected at different time points. Pulmonary nuclear factor kappaB (NFkappaB) activation was investigated with electromobility shift assay. mRNA expression of inflammatory mediators and their corresponding receptors were detected with Ribonuclease Protection Assay. Protein expression was detected by ELISA and western blotting. Inducible NO synthase (iNOS) expression was monitored by RT-PCR and iNOS activity by conversion of l-arginine to citrulline. Immune cells were sampled by bronchoalveolar lavage (BAL) and classified. RESULTS: LPS application induced CD14-, but not TLR4 protein expression in control mice. Activation of pulmonary NFkappaB was observed within 60 min in control, but not in TLR4-D mice. Six hours of LPS administration induced a significant increase in pulmonary tumour necrosis factor alpha-, interleukin-1beta- and interleukin-6 mRNA and protein expression in control mice compared to TLR4-D mice. Furthermore, LPS induced a significantly higher increase of the iNOS expression and catalytic activity in control mice than in TLR4-D mice. BAL revealed an increase in total cell count in all LPS treated mice. CONCLUSION: Our findings suggest that TLR4 plays a key role for regulating the expression of relevant cytokines within the lung during endotoxic shock.

Animals↗

PubMed2006

Early and mid-term results of the Shelhigh stentless bioprosthesis in patients with active infective endocarditis.

AIMS: This study investigated the early and mid-term results following valve replacement with the new Shelhigh stentless bioprosthesis made entirely of biological material in patients with active infective endocarditis (AIE). MATERIAL AND METHODS: Between 02/2000 and 12/2004, 164 patients (n = 122 men, mean age 59, 18-85 years) received implantation of an AIE Shelhigh stentless bioprosthesis in the aortic, mitral, tricuspid or pulmonary position. A total of 119 patients (72.6%) had native AIE and 45 (27.4%) prosthetic AIE. A large proportion of the patients reached the operating room in a condition of cardiac decompensation: 37 (22.6%) patients were intubated, 40 (24.4%) had protracted septic shock and 41 (25.0%) required intensive catecholamine treatment. Surgery was regarded as urgent in 94 patients (57.4%) and was performed as an emergency procedure in 70 (42.6%). The mean follow-up time is 1.5 +/- 0.11 years (range, 5 months to 5.2 years). Echocardiographic follow-up examinations were performed early postoperatively and after 12 months. RESULTS: In terms of the operative indication, we found a highly significant difference in the survival rate between patients who were operated on urgently vs in an emergency. In patients who died within 30 days, the main cause of death was septic multiorgan failure (67.6%). Only three patients required reoperation due to reinfection of the Shelhigh bioprostheses; this represents a reinfection rate of 1.8% in relation to the whole cohort. The postoperative echocardiographic examinations showed the Shelhigh valves to have very good hemodynamics without relevant pressure gradients. CONCLUSION: Our experience in the use of Shelhigh bioprostheses in patients with native and prosthetic endocarditis show the early and mid-term results, in particular the low reinfection rate and the good hemodynamics, to be comparable with the results achieved using homografts. Since these prostheses are readily available and their implantation straightforward, they are increasingly being used in patients with endocarditis. These promising results need to verified in the long term.

Adolescent↗

PubMed2006

Different effects of arborinine alkaloid obtained from Brazilian Erthela baihensis on spleen and thymus cells stimulated in vitro with different mitogens.

The present study has examined the effects of arborinine, an alkaloid obtained from Erthela bahiensis, a Brazilian plant popularly used as diuretic, antidiabetic, antithermic and expectorant, on the viability and function of immune system cells in vitro using a murine model. Rat spleen and thymus cells were cultured with 10 nM, 1 microM, 10 microM of the drug in the presence or absence of pokeweed (PWM), lipopolysaccharide (LPS), or concanavallin (ConA) mitogens. Cellular proliferation was analyzed by H3-thymidin uptake after 48 and 72 hr. Our results showed an inhibitory effect of arborinine on splenocytes proliferation under ConA or PWM stimulation and increased apoptosis on splenocytes and thymocytes stimulated with PWM in 24 hr. A decrease was observed on Interferon gamma (IFN-gamma) production by ConA- or LPS-stimulated splenocytes in 48 hr and 72 hr and ConA- or PWM-stimulated thymocytes in 72 hr. In contrast, an increase on lymphoproliferation was observed on LPS-stimulated splenocytes and ConA- or PWM-stimulated thymocytes in 48 hr. On this period, apoptosis decreased on LPS- or PWM-stimulated splenocytes and IFN-gamma production increased in PWM stimulated thymocytes. Arborinine also induced a decrease on Interleukin-10 production by splenocytes and thymocytes stimulated with ConA or PWM. There was no significant variation on the necrosis rate of the cells treated with arborinine or any change on their viability or function values in the absence of mitogenic stimulus.

Alkaloids↗

PubMed2006

Comparison of microbiological results of needle puncture vs. superficial swab in infected diabetic foot ulcer with osteomyelitis.

AIM: To study prospectively two methods for the bacteriological diagnosis of osteomyelitis related to diabetic foot ulcer: needle puncture performed across normal skin surrounding the foot ulcer and superficial swabbing of the ulcer. PATIENTS AND METHODS: Diabetic patients with a foot ulcer complicated by bone or joint infection, as detected by X-ray imaging, were included in the study. Ulcer swabbing and needle puncture were performed in each patient. To reach the tissue nearest the bone surface, needle puncture was guided by X-ray imaging and the drop of fluid obtained by aspiration was used for both aerobic and anaerobic bacterial culture. RESULTS: Twenty-one diabetic patients were included. The mean number of microorganisms isolated by needle puncture was significantly lower compared with that obtained by superficial swabbing: 1.09 vs. 2.04 (P < 0.02). Three bacterial species were isolated by needle puncture only in one patient while three or more bacterial isolates were obtained by superficial swabbing in six patients. No bacterial isolate was detected in five patients by needle puncture and in two patients by superficial swabbing. Staphylococcus aureus accounted for 70% of cases (seven patients) when a single bacterial species was obtained by needle puncture. After needle puncture, no wound complication or infection was observed. CONCLUSION: Culture of samples obtained by needle puncture revealed one or two bacterial isolates in two-thirds of diabetic patients with osteomyelitis following foot ulcer. Given the lack of complications, this invasive diagnostic technique should be considered for deep direct sampling in diabetic patients with osteomyelitis related to foot ulcer when surgical debridement is contraindicated or delayed.

Adult↗

PubMed2005

Neospora caninum: infection induced IL-10 overexpression in rat astrocytes in vitro.

The effect of Neospora caninum, a parasite that causes abortion and neuromuscular changes, has been investigated on a major population of neural cells, the astrocytes. Highly enriched astroglial primary cultures obtained from neonatal rats were infected after 21 days of culture. Astroglial reactivity, IL-10 and IFN-gamma expression, and cell viability (lactate dehydrogenase activity, metabolization of tetrazolium salt, and trypan blue exclusion assay) have been investigated after 24 and 72 h of infection. Astroglial hypertrophy, gliofilament reorganization, metabolic changes suggesting hypoxia and a strong IL-10 release have been observed in the infected cells. These results show that neural cells are targets for the parasite and that astrocytes may contribute to the CNS immune response to the parasite.

Animals↗

PubMed2005

In vitro IFN-gamma production by goat blood cells after stimulation with somatic and secreted Corynebacterium pseudotuberculosis antigens.

Corynebacterium pseudotuberculosis is the causal agent of caseous lymphadenitis, a chronic illness that attacks goats and sheep characterized by pyogranulomas formation in lymph nodes and organs. Regarding the current knowledge of the pathogenesis of the caseous lymphadenitis, there is evidence that besides the humoral response the induction of a durable cellular response is fundamental for its control. In this sense, research on antigens of C. pseudotuberculosis that are capable to inducing cellular immunity is an important step for the development of diagnosis tests and more efficient vaccines. In the present study, the interferon-gamma production in cultures of whole blood from infected goats stimulated with secreted bacterial antigen or somatic antigen were used to evaluate the cellular response. The results demonstrated a significant difference in the ability of the two antigens to induce a cellular response. That is, IFN-gamma production was high with cells from infected animals in response to the secreted antigen while IFN-gamma production was low when somatic antigen was used. The concomitant use of these antigens with PWM also showed differences. That is, the secreted antigen increased the IFN-gamma production induced by PWM, while the somatic antigen seems not to have altered the response to PWM.

Animals↗

PubMed2005

Phenotype analysis of lymphocytes of workers with chronic benzene poisoning.

Lifetime exposure to benzene is associated to a variety of blood disorders, and except for the risk of cancer, almost nothing is known concerning health impairment in individuals who are no longer exposed. In Brazil, this exposure is one of the serious problems in workplaces, and many workers have been laid off their jobs due to this intoxication, particularly in the State of Bahia, the largest producer of benzene in Latin America, which is the area of this study. From a larger study to describe health effects and genetic polymorphisms among workers with chronic benzene poisoning (CBP), this previous specific investigation analyzes the association between CBP and the pattern of sub-populations of lymphocytes. The study was performed with a CBP group (n=24) and a control group with other occupational diseases (n=24); both were selected at the Workers Health Study Center in the State of Bahia, Brazil. Clinical and epidemiologic variables were collected from medical records and from a detailed questionnaire. The average age was similar in the two groups (51.1 and 50.7, respectively). Analyzing the mean proportions of the sub-populations of lymphocytes, statistically significant differences were found for T cytotoxic cells (TCD8) (27.9; 19.4; p=0.002) and T helper memory cell (CD4CD45RO) (31.2; 37.0; p=0.015), respectively, for the CBP group and control group. These results should be viewed with caution because of the small sample size, but they strengthen a previous impression that workers exposed to benzene have their immune system impaired, even in the long term, which may contribute to some disorders and carcinogenesis process. These workers must be strictly followed up in a medical surveillance program. Although this problem has been known for a long time, this is the first attempt to study these specific effects in Brazil.

Adult↗

PubMed2005

Serologic immunoreactivity to Neospora caninum antigens in dogs determined by indirect immunofluorescence, western blotting and dot-ELISA.

Neospora caninum, is a coccidian protozoan known as a major cause of bovine abortion and canine neuropathies. The aim of the present study was to develop a reliable and quick test to detect antibodies to N. caninum in dog sera. Sixty-five serum samples from dogs, including 35 positive and 30 negative for N. caninum antibodies were used for standardization of the test. In parallel, immunoreactivity of the sera to Toxoplasma gondii antigens was investigated using a passive agglutination test. A dot-ELISA test, using soluble extract of N. caninum tachyzoites on nitrocellulose ester membranes, was developed and standardized. SDS-PAGE and complementary analysis of reactivity by Western blotting were used for the characterization of the immunoreactive fractions of all tested sera. The sensitivity and specificity of the dot-ELISA were 94 and 73%, respectively, compared to IFAT at a cut-off of 1:50, and 87 and 100% compared to IFAT at a cut-off of 1:25. Among the sera that tested positively for both IFAT and dot-ELISA, only 8.6% were reactive to T. gondii. The most immunoreactive fractions in Western blots were the 14-, 33-, 42- and 55 kDa bands, with percentages of 42, 60, 42 and 37%, respectively. The 60 kDa band showed a non-specific reaction in 43% of neosporosis-negative animals by both dot-ELISA and IFAT. These results indicate that the dot-ELISA using N. caninum antigen present good sensitivity and specificity, and might be used as a screening test to detect antibodies to N. caninum in dogs.

Animals↗

PubMed2005

Myocardial viability assessment in patients with highly impaired left ventricular function: comparison of delayed enhancement, dobutamine stress MRI, end-diastolic wall thickness, and TI201-SPECT with functional recovery after revascularization.

This study compared different magnetic resonance imaging (MRI) methods with Tl(201) single photon emission computerized tomography (SPECT) and the "gold standard" for viability assessment, functional recovery after coronary artery bypass grafting (CABG). Twenty patients (64+/-7.3 years) with severely impaired left ventricular function (ejection fraction [EF] 28.6+/-8.7%) underwent MRI and SPECT before and 6 months after CABG. Wall-motion abnormalities were assessed by stress cine MRI using low-dose dobutamine. A segment with a nonreversible defect in Tl(201)-SPECT and a delayed enhancement (DE) in an area >50% of the entire segment, as well as an end-diastolic wall thickness <6 mm, was defined as nonviable. The mean postoperative EF (n=20) improved slightly from 28.6+/-8.7% to 32.2+/-12.4% (not significant). Using the Tl(201)-SPECT as the reference method, end-diastolic wall thickness, MRI-DE, and stress MRI showed high sensitivity of 94%, 93%, and 84%, respectively, but low specificities. Using the recovery of contractile function 6 months after CABG as the gold standard, MRI-DE showed an even higher sensitivity of 99%, end-diastolic wall thickness 96%, stress MRI 88%, and Tl(201)-SPECT 86%. MRI-DE showed advantages compared with the widely used Tl(201)-SPECT and all other MRI methods for predicting myocardial recovery after CABG.

Aged↗

PubMed2005

Primary fibrosarcoma of the liver infiltrating the right atrium of the heart.

A rare case of a primary fibrosarcoma of the liver infiltrating the right heart is described in a 72-year-old woman. The patient presented with a history of progressive dyspnea and ascites and her general condition was poor. Preoperative cardiac magnetic resonance (CMR) imaging revealed a large mass, which originated from the liver and had infiltrated the right atrium via the inferior vena cava. The patient underwent tumor resection yet died shortly afterwards. Histologically the mass was classified as a fibrosarcoma with positive immunostaining for vimentin. We report the CMR imaging characteristics in this uncommon case. Preoperative CMR proved to be useful for clinical decision making and the planning of surgery.

Aged↗

PubMed2004

Identification and quantification of gossypol in cotton by using packed micro-tips columns in combination with HPLC.

Self-packed micro-tip columns containing a C18-bonded silica stationary phase, based on the same principles as solid-phase extraction methods, were used to obtain gossypol and related sesquiterpenoid aldehyde-enriched fractions. The enriched metabolite fractions were then analyzed by optimized high-performance liquid chromatography (HPLC) with a C18 column (4.6 mm x 25 cm) eluted with the binary mobile phase acetonitrile-0.1% aqueous TFA solution (80:20). This method has proven to be highly reproducible. The precision and accuracy, as %RSD and %RME values, were determined to be less than 15% for the method. The minimum detection limit of gossypol was determined to be 10 ng (absolute gossypol). Absolute recovery was greater than 94% with a standard deviation of +/-3.68%. This is a simple, fast, and cost-effective method for isolation, identification, and quantification of gossypol and related secondary metabolites. Comparative analysis of gossypol content was performed on different parts of the cotton plant (seeds, stems and leaves) of two different cultivars of Gossypium hirsutum L. (Acala(1517-70) and OR19). The results indicate that the OR19 cv naturally contains higher gossypol levels than the Acala cv. It was also found that treatment of leaves with a Verticillium dahliae-derived elicitor induced production of deoxyhemigossypol rather than gossypol.

Chromatography, High Pressure Liquid↗

PubMed2004

17 Beta-estradiol regulates nNOS and eNOS activity in the hippocampus.

Estrogen treatment in symptomatic postmenopausal women appears to improve cognitive performance including memory, an effect which may involve enhanced nitric oxide formation in hippocampal neurons. To study whether 17beta-estradiol (E2) affects NO synthase activity in the hippocampus, we investigated the influence of E2 on hippocampal NO synthase expression and activity in female rats. Ovariectomy, which significantly decreased E2 serum levels, reduced neuronal (nNOS) and endothelial NO synthase (eNOS) expression and Ca(2+)-dependent NOS activity. E2 substitution reversed these effects. It is concluded that E2 increases nNOS and eNOS expression and activity in female hippocampus and thus improves hippocampal function.

Animals↗

PubMed2004

Tbx2 represses expression of Connexin43 in osteoblastic-like cells.

Tbx2 belongs to a family of developmental transcription regulatory factors. We evaluated whether the gap junction protein Connexin43 (Cx43), an important regulator of osteoblast function and bone development, may be a downstream target gene regulated by Tbx2. The Cx43 promoter contains direct repeats of the consensus T-box binding motif, TCACAC, and moreover, Tbx2 and Cx43 show overlapping expression domains in precursors to bone and in osteoblasts. In vitro analysis showed that the Cx43 promoter contains two Tbx2 binding sites, and this binding was dependent on the TCACAC consensus sequence. Transient transfection analysis with a Cx43 promoter-driven lacZ reporter construct revealed negative regulation mediated by these two Tbx2 binding sites in osteoblast-like cells. Thus, downregulation of Tbx2 led to de-repression of wild-type Cx43 promoter activity, whereas a promoter construct with mutated binding sites showed no de-repression. In stably transfected osteosarcoma cells in which expression of the endogenous Tbx2 gene was downregulated with a Tbx2 antisense construct, a marked de-repression of the endogenous Cx43 gene was observed. This was accompanied by a marked increase in the abundance of Cx43 gap junctions and increased functional gap junction-mediated cell-cell communication. Analysis of lacZ expression in transgenic mice containing the mutated Cx43 promoter-driven lacZ construct further suggested de-repression of the Cx43 promoter in limb buds, a region destined to give rise to long bones of the limbs. Taken together, these findings indicate that the promoter of Cx43 is repressible by Tbx2, both in cultured osteoblast-like cells in vitro and likely in the developing embryo.

Animals↗

PubMed2004

Orthotopic heart transplantation with concurrent coronary artery bypass grafting or previous stent implantation.

We describe three cases of donor hearts with preexisting coronary artery disease already diagnosed prior to transplantation: two were treated by coronary artery bypass grafting during the transplant procedure and one by angioplasty with stenting during the donor screening angiography. All three donor organs would otherwise have been rejected, depriving potential recipients of organ transplantation. All patients had an uneventful post-operative course with follow-up completed 22, 40 and 43 months after orthotopic transplantation showing patency of the stent and bypass grafts in the early (1 and 9 months) and late (22, 24 and 37 months) coronary angiography. Our results suggest that in this era of acute organ shortage donor hearts requiring bypass or stenting, which form a small but significant donor subgroup, can be used effectively and safely when matched to the recipients' age and medical condition.

Aged↗

PubMed2004

Three-phase partitioning as an efficient method for extraction/concentration of immunoreactive excreted-secreted proteins of Corynebacterium pseudotuberculosis.

The three-phase partitioning (TPP) technique was used upstream to isolate/concentrate secreted proteins from Corynebacterium pseudotuberculosis cultured in a complex liquid medium. Several parameters of the TPP technique (15, 30, or 60% ammonium sulfate concentration; 4.0, 5.5, or 7.0 pH; and primary (n) or tertiary (t)-butanol solvent isomer) were varied to determine the optimal recovery of serologically and cellularly immunoreactive extracted proteins. A TPP extraction made with 30% ammonium sulfate and an initial pH of 4.0 gave the best humoral and cellular immunoreactivity of caseous lymphadenitis infected goats. In particular, two immunogenic secreted (16 and 125 kDa) proteins, which had not been found by other extraction methods, were identified.

Ammonium Sulfate↗

PubMed2004

Role of B cells and macrophages in microvascular disease after heart transplantation.

BACKGROUND: The role of B cells and macrophages in microvascular disease after heart transplantation remains controversial. METHODS: Out of a total of 809 endomyocardial biopsies without evidence of acute cellular rejection (n = 422, 72 females and 350 males, median age 46 years), 393 without evidence of the quilty phenomenon were investigated zero to ten years after heart transplantation. Vascular reaction (endothelial cell swelling and vessel wall thickening) was graded by H&E staining, and immunohistochemistry was performed for T cells (clone UCHL1), B cells (clone L26) and macrophages (clone KP1) and evaluated semi-quantitatively (light microscopy x 200). RESULTS: Positive reaction for T cells and macrophages as well as evidence of endothelial cell swelling decreased with time after heart transplantation. Positive reactions for B cells were less frequent and increased slightly during the observation time, while vessel wall thickening dominated the last observation interval between the fourth and tenth years. Severity of vascular reaction was independent of immunohistochemical evidence of B cells or macrophages. CONCLUSIONS: While activation of the humoral and the non-specific immunological system is common after heart transplantation, microvascular alterations seem to develop independently of these findings.

Adult↗