Mechanisms of edema formation in experimental autoimmune encephalomyelitis. The contribution of inflammatory cells.

Most of the central nervous system (CNS) endothelium regulates the passage of solutes and functions as a blood-brain barrier (BBB). During experimental autoimmune encephalomyelitis (EAE), an inflammatory demyelinating disease of the CNS, loss of BBB function occurs. The authors have previously shown an increase in endothelial transcytotic activity associated with decreased mitochondrial content as evidence of BBB dysfunction in EAE. These changes occurred in the capillary bed and correlated with CNS edema and clinical signs. In the present report, a fixation procedure before infusion of the intravascular tracer horseradish peroxidase (HRP) in rats at the height of clinical EAE has been used. In these animals, tracer leakage was only noted in inflamed venules with diameters of 12 to 19 mu. The authors detected several mechanisms of passive leakage: 1) increased junctional permeability; 2) increased interendothelial space; 3) leakage alongside migrating inflammatory cells. Some small capillaries showed necrotic changes with minimal tracer leakage. This report demonstrates that BBB disruption also occurs via nonendocytic mechanisms that may be induced by inflammatory cells.     

Epidemiological Investigation of Vaginal Saccharomyces cerevisiae Isolates by a Genotypic Method

Saccharomyces cerevisiae is a ubiquitous, ascomycetous yeast, and vaginitis caused by this organism has been reported only very rarely. The aim of the present investigation was to assess the epidemiological relatedness of a group of vaginal and commercial S. cerevisiae isolates by a previously reported genetic typing method, which divided the isolates into two broad groups with numerous subtypes. Nineteen S. cerevisiae isolates obtained from patients suffering from vaginitis and four isolates from commercial products in the same city were analyzed. The cellular DNA from each isolate was digested with the restriction endonuclease EcoRI, and restriction fragment length polymorphisms were generated by horizontal gel electrophoresis. The results showed that although vaginal isolates did not cluster in any particular genetic subtype, multiple patients were infected with indistinguishable strains (there were nine distinct strains among 23 isolates). For two of three patients, all three with two episodes of S. cerevisiae vaginitis, different strains were isolated during the recurrence of this disease. Three other patients with indistinguishable isolates were epidemiologically related in that two were practitioners in the same clinic and the third was a patient at this clinic. We also found that one commercial strain was indistinguishable from the strain isolated from three different women at the time that they were suffering from vaginitis. The findings of the present study suggest that some S. cerevisiae strains may possess properties permitting persistence in the human host. Furthermore, person-to-person contact and the proliferation of the use of S. cerevisiae as a health-food product, in home baking, and in home brewing may be a contributing factor in human colonization and infection with this organism.     

Prostate cancer is part of the hereditary non-polyposis colorectal cancer (HNPCC) tumor spectrum

The recognized urologic tumor spectrum in hereditary non-polyposis colon cancer includes ureteral and renal pelvis malignancies. Here, we report a family in which the proband, who had three metachronous adenocarcinomas of the colon and rectum (at ages 54, 57, and 60), presented with an adenocarcinoma of the prostate at age 61. Immunohistochemical (IHC) staining of colonic, rectal, and prostatic tumor tissues demonstrated lack of expression of both MSH2 and MSH6. Accordingly, microsatellite instability (MSI) was found in the rectal, colonic, and prostatic tumors. The kindred complies with the Amsterdam criteria for HNPCC, as five members over three generations had colorectal cancer. Molecular investigations were initiated when the proband's son presented with an adenocarcinoma of the colon at age 35. Southern blotting analysis of genomic DNA led to identification of a novel genomic deletion encompassing exon 5 of the MSH2 gene. Although prostate cancer has occasionally been described in HNPCC families, to the best of our knowledge, this is the first report where the MSI and IHC analysis of the prostatic adenomcarcinoma clearly link its aetiology to the germline mismatch repair mutation. Hence, prostate cancer should be included in the HNPCC tumor spectrum.     

Epidemiology of extended-spectrum beta-lactamase-producing Enterobacter isolates in a Spanish hospital during a 12-year period

Fifteen Enterobacter clinical isolates (11 Enterobacter cloacae isolates, 3 Enterobacter aerogenes isolates, and 1 Enterobacter gergoviae isolate), representing 0.4% of all Enterobacter isolates recovered in our hospital from 1989 to 2000, were suspected of harboring an extended-spectrum beta-lactamase (ESBL). These isolates were recovered from 14 different patients. ESBLs were transferred by conjugation into an Escherichia coli recipient strain. Pulsed-field gel electrophoresis (PFGE) revealed a single clone of E. aerogenes and six different clones of E. cloacae. Four of these E. cloacae clonal types were represented by only one isolate each, but the other two were represented by three and four isolates, respectively. Isoelectric focusing, susceptibility phenotyping, PCR analysis, and sequencing demonstrated the presence of three different ESBLs. The most frequent was the recently characterized CTX-M-10 ESBL, which was found in the E. gergoviae isolate and in all but one of the E. cloacae isolates. The remaining E. cloacae isolate harbored a TEM-27 ESBL, and the three E. aerogenes isolates harbored a TEM-24 ESBL. PFGE revealed that our E. aerogenes strain was indistinguishable from the French TEM-24-producing E. aerogenes endemic clone. Although a low prevalence of ESBL-producing Enterobacter isolates was found in our institution over a 12-year period, a diversity of nonepidemic E. cloacae clones was detected, as was the persistence of the CTX-M-10 beta-lactamase. The presence of the TEM-24-producing E. aerogenes French clone in our institution also demonstrates the intercountry dissemination of ESBL-producing isolates.     

T cell subset alterations in idiopathic glomerulonephritis

Peripheral blood lymphocytes from 15 healthy controls and 59 patients with idiopathic glomerulonephritis were studied to determine whether an imbalance exists among human T cell subsets in these diseases. Twenty of the patients studied had a minimal change nephropathy (10 with nephrotic syndrome and 10 in sustained remission); 27 had a membranous glomerulonephritis (12 with nephrotic syndrome, six with isolated proteinuria and nine in complete remission); 12 patients had an IgA glomerulonephritis with heamaturia and mild proteinuria. Monoclonal antibodies directed at human T lymphocyte subsets termed OKT3, OKT4 and OKT8 were used in an indirect immunofluorescence assay in all cases. Patients with minimal change nephropathy, with or without nephrotic syndrome and patients with IgA glomerulonephritis showed mean values of OKT3⁺ cells (total peripheral T cells), helper OKT4⁺ cells, suppressor OKT8⁺ cells and OKT4⁺/OKT8⁺ cell ratio, in the normal range. Only the group of patients with membranous glomerulonephritis and nephrotic syndrome presented a mean OKT4⁺/OKT8⁺ ratio greater than the normal group (percentages: 2·43±0·3 vs 1·6±0·1 s.e.m.; P<0·02). This increased ratio was due to a reduction in the OKT8⁺ cell subset compared to the healthy subjects (percentages: 27·6±2·9 vs 36·8±1·4 s.e.m.; P<0·01). Our data shows that the functional lymphocyte disorders previously described in minimal change nephropathy and IgA glomerulonephritis are not due to a numerical imbalance of lymphocyte subsets. Such an imbalance of lymphocyte subsets was specifically observed in membranous glomerulonephritis with nephrotic syndrome. The true significance of this finding has to be clarified by longitudinal studies and functional tests.     

Antidepressant response and well-being in pre-, peri- and postmenopausal women with major depressive disorder treated with fluoxetine

BACKGROUND: We assessed the impact of menopausal status on treatment response and well-being in a cohort of outpatient women with major depressive disorder (DSM-III-R criteria), who received treatment with fluoxetine (20 mg/day for 8 weeks). METHODS: Menopausal status was defined based on age, presence of menstrual irregularity or amenorrhea and vasomotor symptoms. Remission and response of depression were defined as a 17-item Hamilton Depression Rating Scale (HAM-D-17) score or=50%, respectively. Well-being was assessed by self-rating with the Symptom Questionnaire. Remitters were followed up for 28 additional weeks. RESULTS: No differences in rates of response and remission as well as in levels of well-being were observed among pre- (n = 121), peri- (n = 28) and postmenopausal (n = 35) women at the endpoint of the acute phase, even after adjustment for baseline depression severity. Residual symptoms, however, were significantly more common in postmenopausal women, except for the continuation phase endpoint. Differences in residual symptoms during the acute phase subsided after adjustment for baseline depression severity. CONCLUSIONS: Overall, menopausal status did not significantly affect the response to fluoxetine treatment and the degree of posttreatment well-being among major depressive disorder patients.     

Ultrasound-guided in utero injections allow studies of the development and function of the eye

Ultrasound-guided in utero injections into the brain of murine embryos has been shown to facilitate gene delivery. We investigated whether these methods would allow gene transfer into ocular structures. Gene transfer using retroviral vectors or electroporation was found to be quite effective. We determined the window of time, as well as compared several strains of mice, that yield a high degree of survival and successful gene transfer. Several retroviral constructs were tested for expression and coexpresssion of two genes in retinal cell types. In addition, a retroviral vector was engineered to give cone photoreceptor-enriched expression, and a retroviral vector was demonstrated to provide RNAi-mediated loss-of-function. These methods enable access to early ocular structures and provide a more rapid method of assessment of gene and promoter function than possible using genetically engineered mice.     

Immunosuppressive effects of (2R,5R)-6-heptyne-2,5-diamine an inhibitor of polyamine synthesis: I. Effects on mitogen-induced immunoglobulin production in human cultured lymphocytes.

The consequences of specific inhibition of polyamine biosynthesis by (2R,5R)-6-heptyne-2,5-diamine (MAP) a potent inhibitor of L-ornithine decarboxylase (ODC), on immunoglobulin (Ig) production were studied in cultured human peripheral blood lymphocytes stimulated with pokeweed mitogen (PWM). MAP inhibits the usual PWM-induced increase of polyamine (putrescine, spermidine and spermine) concentrations and reduces concomitantly cell replication. In parallel with these biochemical effects, IgG and IgM production are diminished, a 95% decrease being observed at 100 microM MAP concentration. That the suppressive effects of the ODC inhibitor result from polyamine deficiency, and not from unrelated pharmacological effects, is demonstrated by the restoration of normal Ig production when 10 microM putrescine or spermidine are added to the culture medium. These findings established that the cellular immunological response can be affected by specific inhibition of polyamine biosynthesis and deserve further consideration both under in vitro and in vivo conditions.     

Hypomethylation of the human HLA-DRα gene in breast carcinomas and autologous metastases

The methylation pattern of the human HLA-DR gene was analyzed in normal breast tissues, breast primary tumors and lymphonodal metastases isolated from patients carrying breast carcinomas. In breast adenomas and also in normal tissues (including breast, muscle, brain, sperm and T- and B-lymphocytes), the HLA-DR gene is hypermethylated at the CCGG and GCGC sites. In all tissues studied, the only constantly unmethylated region is located in the 5 portion of the gene, near the promoter sequence. Further, the results indicate that the HLA-DR gene is hypomethylated in carcinomas and in the relative metastatic lymph nodes. It is suggested that hypomethylation of the human HLA-DR gene could be proposed as a molecular marker of malignant breast tumors.     

Absence of TREM2 polymorphisms in patients with Alzheimer's disease and Frontotemporal Lobar Degeneration

Triggering Receptor Expressed on Myeloid cells (TREM)2 deficiency originates a genetic syndrome characterized by bone cysts and presenile dementia, named Nasu-Hakola disease (NHD). Early onset dementia and marked involvement of frontal regions are features characterizing both NHD and other kinds of neurodegenerative disorders, such as Frontotemporal Lobar Degeneration (FTLD), and, in some cases, Alzheimer's disease (AD). Three Single Nucleotide Polymorphisms (SNPs) in TREM2 coding region were screened by allelic discrimination in a population of probable AD patients as well as FTLD patients as compared with age-matched controls. In addition, mutation scanning of the coding region of TREM2 gene was carried out in 7 patients with early onset AD (EOAD), 16 FTLD, and 20 controls. None of the SNPs analyzed was present, either in patients or controls. Moreover, mutation scanning of the five exons of TREM2 failed to detect the presence of novel polymorphisms. These data demonstrate that TREM2 coding region is highly conserved, implying a crucial role of this receptor. Further studies, including a functional analysis, are certainly required to clarify the role of TREM2 in neurodegenerative processes.