Vaccinia virus encodes a protein with similarity to glutaredoxins

Recently, we have reported the complete nucleotide sequence of vaccinia virus (Goebel, S. J., Johnson, G. P., Perkus, M. E., Davis, S. W., Winslow, J. P., and Paoletti, E. 1990, Virology 179, 247-266). Approximately 2.2 kbp leftward of the large subunit of ribonucleotide reductase resides a 108-amino acid open reading frame, O2L (nt 62,851-62,528) with significant similarity to known glutaredoxins. The deduced amino acid sequence of open reading frame O2L is 28.7% identical to the yeast and Escherichia coli proteins and greater than 40% identical to various mammalian glutaredoxins. Similar patterns of hydrophobicity as well as alpha-helix and beta-sheet potentials suggest that O2L and the glutaredoxins share a similar secondary structure. Furthermore, a common function is inferred by the presence of a highly conserved redox-active site.     

Infection of the Laboratory Mouse with the Intracellular Pathogen Ehrlichia chaffeensis

To determine the basis of susceptibility and resistance to human monocytic ehrlichiosis (HME), immunocompetent and immunocompromised mice were infected with Ehrlichia chaffeensis and bacterial loads were measured by PCR and by immunohistochemistry. Immunocompetent (C.B-17 and C57BL/6) mice cleared the bacteria within 10 days, but immunocompromised SCID and SCID/BEIGE mice developed persistent infection in the spleen, liver, peritoneal cavity, brain, lung, and bone marrow and became moribund within 24 days. Both immunocompromised strains lack T and B lymphocytes, but the SCID/BEIGE strain is also deficient in natural killer (NK) cell function. During advanced stages of disease, the infections were associated with wasting, splenomegaly, lymphadenopathy, liver granulomas and necroses, intravascular coagulation, and granulomatous inflammation. Histochemical and immunohistochemical localization studies confirmed the presence of bacteria in tissues, and viable bacteria were cultured from infected animals. The data reveal that T and/or B cells play an essential role during resistance of immunocompetent mice to infection with E. chaffeensis and demonstrate the utility of immunocompromised mice as an experimental model for the study of HME.     

Elevated production of interferon-gamma and interleukin 4 by mature T cells from autoimmune lpr mice correlates with Pgp-1 (CD44) expression

The cell surface glycoprotein, Pgp-1 (CD44), has been shown to be a marker of murine memory T lymphocytes. When activated, Pgp-1hi memory T cells produce strikingly higher amounts of interferon-gamma (IFN-gamma) than naive Pgp-1lo T cells, yet both subsets make similar levels of interleukin (IL)2. Whereas Pgp-1hi cells represent only 20%-25% of peripheral T cells from most strains, this marker is expressed by the vast majority (greater than 90%) of T cells from autoimmune MRL mice homozygous for the lymphoproliferation (lpr) gene. The massive lymphadenopathy that develops in lpr/lpr mice is composed of both non-mature (CD4-CD8-) T cells as well as a greatly expanded number (up to 300-fold) of mature (CD4+CD8-,CD4-CD8+) T cells. Paralleling the expression of high levels of Pgp-1, we find that compared to normal mouse T cells, the lpr mature T lymphocyte subsets are also very high producers on a per cell basis of IFN-gamma and, for the CD4+ subset, IL 4. Increased concentrations of IFN-gamma and IL 4 produced by large numbers of lpr Pgp-1hi mature T cells could contribute to the autoimmune syndrome in MRL lpr/lpr mice through the effects of these cytokines on augmenting MHC class II expression and production of certain classes of antibodies.     

Enhanced molecular volume of conservatively pegylated Hb: (SP-PEG5K)6-HbA is non-hypertensive

Recent studies have suggested that the "pressor effect" of acellular Hb is a consequence of perturbation of the macro-and microcirculatory system in multiple ways, and that PEGylation is an effective approach for controlling the same. In an attempt to confirm this concept, a new and simple thiolation mediated, maleimide chemistry-based conservative PEGylation protocol has been developed to conjugate multiple copies of PEG-chains to Hb. This approach combines the high reactivity of maleimides towards thiols with the propensity of iminothiolane to derivatize the epsilon-amino groups of proteins into reactive thiol groups, with conservation of their positive charge. One of the PEGylated products, namely (SP-PEG5K)6-HbA, that carries on an average six copies of PEG5000 chains per Hb, is non-hypertensive in hamster top load and in rat 50% exchange transfusion models. This hexa-PEGylated-Hb has (i) a hydrodynamic volume corresponding to that of an oligomerized Hb of 256kDa, (ii) a molecular radius of approximately 6.8 nm, (iii) high oxygen affinity, (iv) lowered Bohr effect, and (v) increased viscosity and colloidal osmotic pressure. These properties of (SP-PEG5K)6-HbA are consistent with the emerging new paradigms for the design of Hb based oxygen carriers and confirm the concept that the "pressor effect" of Hb is a multifactorial event. The thiolation mediated maleimide chemistry-based PEGylation protocol described here for the generation of (SP-PEG5K)6-Hb is simple, highly efficient, and is carried out under oxy conditions. The results demonstrate that a non-hypertensive PEG-Hb can be generated by conjugation of a lower number of PEG chains than previously reported.     

Immunological parameters in girls with Turner syndrome

Disturbances in the immune system has been described in Turner syndrome, with an association to low levels of IgG and IgM and decreased levels of T- and B-lymphocytes. Also different autoimmune diseases have been connected to Turner syndrome (45, X), thyroiditis being the most common.     

A point mutation in the 5' splice site of the dystrophin gene first intron responsible for X-linked dilated cardiomyopathy

X-linked dilated cardiomyopathy (XLDC) is a familial heart disease presenting in young males as a rapidly progressive congestive heart failure, without clinical signs of skeletal myopathy. This condition has recently been linked to the dystrophin gene in some families and deletions encompassing the genomic region coding for the first muscle exon have been detected. In order to identify the defect responsible for this disease at the molecular level and to understand the reasons for the selective heart involvement, a family with a severe form of XLDC was studied. In the affected members, no deletions of the dystrophin gene were observed. Analysis of the muscle promoter, first exon and intron regions revealed the presence of a single point mutation at the first exon-intron boundary, inactivating the universally conserved 5' splice site consensus sequence of the first intron. This mutation introduced a new restriction site for MseI, which cosegregates with the disease in the analyzed family. Expression of the major dystrophin mRNA isoforms (from the muscle-, brain- and Purkinje cell-promoters) was completely abolished in the myocardium, while the brain- and Purkinje cell- (but not the muscle-) isoforms were detectable in the skeletal muscle. Immunocytochemical studies with anti- dystrophin antibodies showed that the protein was reduced in quantity but normally distributed in the skeletal muscle, while it was undetectable in the cardiac muscle. These findings indicate that expression of the muscle dystrophin isoform is critical for myocardial function and suggest that selective heart involvement in dystrophin- linked dilated cardiomyopathy is related to the absence, in the heart, of a compensatory expression of dystrophin from alternative promoters.      

Single-stage methods of hypospadias repair

Summary The repair of all hypospadias in one stage represents state of the art in virtually all instances. Implicit in the repair of hypospadias is the correction of chordee, the construction of the neourethra such that the meatus is located on the tip of the glans, and the creation of a penis which is cosmetically appropriate. Described in this paper are most of the currently employed methods utilized for the repair of hypospadias in a single stage. The techniques described herein, in skilled hands, yield good results in 85–90% of cases. Repairs of hypospadias which have been designed and which are recommended for the occasional operator have no place in today's surgical therapy.     

Predictors of Caregiver Participation in an Engagement Strategy to Increase Initiation into a Family-Based Preventive Intervention

Previous studies have shown that engagement strategies can help increase enrollment and initiation of families in evidence-based preventive programs under natural service delivery settings. However, little is known about factors that predict completion of these engagement strategies. This study aimed to examine predictors (i.e., perceived need, perceived barriers, and sociocultural context) of caregiver participation in an evidence-based engagement call strategy. This call was expected to increase initiation into a school-based, family-focused prevention program. In addition, this study examined engagement call completion as a predictor of program initiation among already enrolled families. Participants included ethnically diverse families recruited from three Title I schools (n = 413) who were randomized to receive the prevention program. Results showed that interparental conflict—an indicator of perceived need—was associated with an increased likelihood of completing the engagement call. Furthermore, caregivers from low-socioeconomic status (SES), foreign-born, Spanish-speaking, Hispanic families were more likely to complete the call relative to those from low- and mid-SES, US born, English-speaking, ethnically diverse families. Importantly, engagement call completion was associated with an increased likelihood of program initiation. These findings provide limited support that families with higher perceived needs are more likely to participate in an evidence-based engagement call strategy. Results suggested that the call strategy provides a promising way to reduce attrition from family prevention programs, which is commonly observed between enrollment and initiation. Project Number: R01 DA035855; Date of Registration: 06/15/2014.