1141154113Expression of natural cytotoxicity receptors in peripheral blood NK cell subsets of women with recurrent spontaneous abortions (RSA) or implantation failures

Aim:  To determine if IgA is required for protection against Chlamydia infection in the male reproductive tract (MRT).
Materials and Methods:  Male polyimmunoglobulin receptor knockout mice (PIgR^-/-) and wild-type C57BL/6 (WT) mice were immunised intranasally with chlamydial major outer membrane protein (MOMP) and cholera toxin (CT). MOMP-specific IgG and IgA in serum and prostatic fluids were measured by ELISA. Serum and PF were also assayed for inhibition of in vitro chlamydial infection. Immunized WT and PIgR^-/- mice were challenged by direct inoculation of C. muridarum into the meatus urethra. Four weeks post challenge Chlamydia levels in the penile urethra, epididymis and testis were determined by PCR.
Results:  Equivalent levels of IgG were found in the serum of both WT and PIgR^-/- mice however IgA in serum of PIgR^-/- mice was 19- to 20-fold higher than in WT animals consistent with the lack of the PIgR IgA transport molecule. IgA levels were significantly lower in PIgR^-/- PF compared to WT PF after both immunization and infection. Only PF from WT but not PIgR^-/- animals was able to inhibit in vitro chlamydial infection. Following challenge significantly higher levels of Chlamydia were recovered from the MRT of PIgR^-/- mice compared to WT animals.
Conclusions:  Male mice lacking a functional PIgR were unable to clear a genital tract Chlamydia infection despite high levels of serum IgA. These data show that mucosal IgA plays a major role in preventing chlamydial infection of the male genital tract and suggest that immunization strategies to protect males should target a strong mucosal IgA response.     

DNA methylation studies on imprinted loci in a male monozygotic twin pair discordant for Beckwith–Wiedemann syndrome

Tierling S, Souren NY, Reither S, Zang KD, Meng‐Hentschel J, Leitner D, Oehl‐Jaschkowitz B, Walter J. DNA methylation studies on imprinted loci in a male monozygotic twin pair discordant for Beckwith–Wiedemann syndrome. Beckwith–Wiedemann syndrome (BWS) is one of the most prevalent congenital disorders predominantly caused by epigenetic alterations. Here we present an extensive case study of a monozygotic monochorionic male twin pair discordant for BWS. Our analysis allows to correlate BWS symptoms, like a protruding tongue, indented ears and transient neonatal hypoglycaemia, to an abnormal methylation at the KvDMR1. DNAs extracted from peripheral blood, skin fibroblasts, saliva and buccal swab of both twins, their sister and parents were analysed at 11 differentially methylated regions (DMRs) including all four relevant DMRs of the BWS region. The KvDMR1 was exclusively found to be hypomethylated in all cell types of the affected BWS twin, while the unaffected twin and the relatives showed normal methylation in fibroblasts, buccal swab and saliva DNA. Interestingly, the twins share a common blood‐specific hypomethylation phenotype most probably caused by a feto‐fetal transfusion between both twins. Because microsatellite analysis furthermore revealed a normal biparental karyotype for chromosome 11, our results point to an exclusive correlation of the observed BWS symptoms to locally restricted epimutations at the KvDMR1 of the maternal chromosome.     

Regeneration and tolerance factor: a correlate of human immunodeficiency virus-associated T-cell activation.

Human immunodeficiency virus (HIV) infection causes extensive phenotypic alterations in lymphocytes. Cellular markers that are normally absent or expressed at low levels on quiescent cells are upregulated throughout the disease course. The transmembrane form of regeneration and tolerance factor (RTF) is expressed at negligible levels on resting T cells but is quickly upregulated following in vitro stimulation and activation. Recently, we reported that expression of RTF was significantly higher in cells from HIV-seropositive (HIV(+)) individuals than in cells from HIV-seronegative (HIV(-)) individuals. Because T cells from HIV(+) individuals express markers reflecting chronic activation, we hypothesized that these in vivo-activated cells would coexpress RTF. Flow cytometry was used to assess RTF expression on activated (CD38(+) and HLA-DR(+)) CD4(+) and CD8(+) T cells. HIV(+) individuals had higher percentages of RTF(+) CD38(+) (P < 0.0001) or RTF(+) HLA-DR(+) (P = 0.0001) CD4(+) T cells than HIV(-) individuals. In HIV(+) individuals, increased percentages of CD4(+) T cells that were RTF(+), RTF(+) CD38(+), and RTF(+) HLA-DR(+) correlated inversely with the absolute number and percentage of CD4(+) T cells and correlated positively with plasma beta(2)-microglobulin concentrations. HIV(+) individuals had higher percentages of CD8(+) T cells that were RTF(+) CD38(+) (P = 0.0001) or RTF(+) HLA-DR(+) (P = 0.0010). In HIV(+) individuals, increased percentages of CD8(+) T cells that were RTF(+) HLA-DR(+) correlated inversely with the percentage of CD4(+) T cells, and high percentages of CD8(+) T cells that were RTF(+) CD38(+) correlated positively with plasma beta(2)-microglobulin levels. These findings strongly suggest that increased RTF expression is a correlate of HIV-associated immune system activation.     

Image-directed percutaneous biopsies with a biopsy gun

Core tissue for histologic study is believed by many pathologists to be more diagnostic than material from needle aspiration. Recently, a biopsy "gun" has been introduced, which simplifies core biopsies. With this device, 182 biopsies of multiple anatomic sites were performed with ultrasonic, computed tomographic, and fluoroscopic guidance and 18-gauge needles. High-quality histopathologic specimens were obtained in 177 of the biopsies, and diagnostic target tissue was obtained in 167. Only three significant complications occurred: one bleeding complication that required transfusion and two cases of pneumothorax that necessitated placement of chest tubes. The biopsy gun eliminated the disjointed movements of conventional "skinny" needle biopsies, and none of the samples demonstrated significant "crush" artifact or obscuring blood, problems that are commonly associated with manual biopsy techniques. Patient discomfort was decreased with this system compared with that of manual biopsies, and the total procedure time was reduced. Because of these distinct advantages, the authors now use the biopsy gun exclusively for all percutaneous biopsies and recommend that other institutions consider the use of this biopsy method.     

[11C]5-HTP and microPET are not suitable for pharmacodynamic studies in the rodent brain

The PET tracer [¹¹C]5-hydroxytryptophan ([¹¹C]5-HTP), which is converted to [¹¹C]5-hydroxytryptamine ([¹¹C]5-HT) by aromatic amino acid decarboxylase (AADC), is thought to measure 5-HT synthesis rates. But can we measure these synthesis rates by kinetic modeling of [¹¹C]5-HTP in rat? Male rats were scanned with [¹¹C]5-HTP (60 minutes) after different treatments. Scans included arterial blood sampling and metabolite analysis. 5-HT synthesis rates were calculated by a two-tissue compartment model (2TCM) with irreversible tracer trapping or Patlak analysis. Carbidopa (inhibitor peripheral AADC) dose-dependently increased [¹¹C]5-HTP brain uptake, but did not influence 2TCM parameters. Therefore, 10 mg/kg carbidopa was applied in all subsequent study groups. These groups included treatment with NSD 1015 (general AADC inhibitor) or p-chlorophenylalanine (PCPA, inhibitor of tryptophan hydroxylase, TPH). In addition, the effect of a low-tryptophan (Trp) diet was investigated. NSD 1015 or Trp depletion did not affect any model parameters, but PCPA reduced [¹¹C]5-HTP uptake, and the k₃. This was unexpected as NSD 1015 directly inhibits the enzyme converting [¹¹C]5-HTP to [¹¹C]5-HT, suggesting that trapping of radioactivity does not distinguish between parent tracer and its metabolites. As different results have been acquired in monkeys and humans, [¹¹C]5-HTP-PET may be suitable for measuring 5-HT synthesis in primates, but not in rodents.     

高效液相色谱流通池法测定紫草中假紫草素的含量

高效液相色谱法测定紫草提取物中的假紫草素时,出现20个峰,需时75min,主峰保留时间55min,常规方法制备工作曲线很费时。本文报道一种新的工作曲线法——流通池法的原理和方法,测定结果与常规方法一致,可以节省时间。     

In situ hybridization for detection of nocardial 16S rRNA: reactivity within intracellular inclusions in experimentally infected cynomolgus monkeys--and in Lewy body-containing human brain specimens

Our previous studies found that experimental infection of BALB/c mice with the Gram-positive bacterium Nocardia asteroides induced a parkinsonian-type syndrome with levodopa-responsive movement abnormalities, loss of nigrostriatal dopaminergic neurons, depletion of striatal dopamine, and intraneuronal inclusions in the substantia nigra (SN) with an appearance similar to Lewy bodies. In the present study, an in situ hybridization technique was developed to detect nocardial 16S ribosomal RNA (rRNA), using a Nocardia-specific probe (B77). Cerebral cortical specimens from cynomolgus monkeys were examined for the presence of nocardial RNA 48 h, 3.5 months, and 1 year after experimental infection with N. asteroides. Hybridization reactions were detected within Nocardia-like structures 48 h after infection and within intracellular inclusion bodies (immunoreactive for α-synuclein and ubiquitin) in one of two 3.5-month-infected monkeys. The in situ hybridization procedure was then applied in a blinded fashion to 24 human SN specimens with Lewy bodies and 11 human SN specimens without Lewy bodies (including five normal controls). Hybridization reactions were detected in nine Lewy body-containing specimens and none of the others. Reactivity was limited to inclusions with the appearance of Lewy bodies, with the exception of one specimen in which intracellular reactivity was also observed in Nocardia-like structures. These results suggest a possible association between Nocardia and neurodegenerative disorders in which Lewy bodies are present.     

Regeneration and tolerance factor modulates the effect of adenosine triphosphate-induced interleukin 1 beta secretion in human macrophages

These studies characterize a molecule known as regeneration and tolerance factor (RTF), which controls inflammation by regulating interleukin 1 beta (IL-1 beta) secretion. Recently, it has been demonstrated that the interaction of adenosine triphosphate (ATP) with the P2X7 purinoceptor induces the secretion of IL-1 beta and initiates the inflammatory response. In these experiments, that the addition of ATP to macrophages was found to induce P2X7 activation and secretion of IL-1 beta. This secretion is enhanced with anti-RTF antibody in combination with exogenous ATP (p< 0.005). RTF is also revealed to be able to influence surface ATPase activity and, increase PI incorporation, which is an indicator of P2X7 activation. We demonstrate that RTF has a role in controlling IL-1 beta secretion by regulating P2X7 activity.