The effect of cannabinoid receptor agonist WIN 55,212–2 on anxiety‐like behavior and locomotion in a genetic model of absence seizures in the elevated plus‐maze

GAERS and NEC rats were treated with cannabinoid 1/2 receptor agonist WIN 55,212‐2 2 mg/kg and tested on the Elevated Plus‐Maze.     

Effects of between generations changes in nutrition type on vaginal smear and serum lipids in Sprague–Dawley rats

Objective: Previous studies established that between generations changes in feeding protocol can have significant impact on reproductive physiology. The aim of the study was to determinate effects of mothers' nutrition and nutrition of the offspring on the characteristics of vaginal smear and serum lipid content.

Methods: Ten female rats were randomly divided in two groups; first group fed with food containing high content of saturated fatty acids (HFD) and the second with standard laboratory chow (CD). After coupling and lactation period their offspring were further randomly divided into two subgroups fed HFD or CD forming four study groups: (a) CD–CD, (b) CD–HFD, (c) HFD–CD and (d) HFD–HFD. The dams and offspring at the age of 37 and 18 weeks, respectively, were subjected to biochemical analysis of the blood and cytological analysis of the vaginal smears. Additionally body weight was recorded and body mass index (BMI) was calculated.

Results: The HFD–HFD group presented with highest levels of triglycerides and the CD–HFD with the highest levels of cholesterol. Therefore, triglyceride and cholesterol levels were significantly different among the groups (p?=?0.001 and p?=?0.002, respectively). Vaginal cytological smears analysis showed features of irregular phase interchanges or extended estrous phase in offspring of high-fat fed dams.

Conclusion: Maternal HFD consumption predisposes offspring to increased risk of developing metabolic abnormalities and estrous disorders.     

Antibodies that bind complex glycosaminoglycans accumulate in the Golgi

Light (L) chains that edit anti-DNA heavy (H) chains rescue B-cell development by suppressing DNA binding. However, exceptional editor L chains allow B cells to reach splenic compartments even though their B-cell receptors remain autoreactive. Such incompletely edited B cells express multireactive antibodies that accumulate in the Golgi and are released as insoluble, amyloid-like immune complexes. Here, we examine examples of incomplete editing from the analysis of variable to joining (VJ) gene junction of the variable (Vλx) editor L chain. When paired with the anti-DNA heavy chain, V_H56R, the Vλx variants yield antibodies with differing specificities, including glycosaminoglycan reactivity. Our results implicate these specificities in the evasion of receptor editing through intracellular sequestration of IgM and the release of insoluble IgM complexes. Our findings can be extrapolated to human L chains and have implications for understanding a latent component of the Ig repertoire that could exert pathogenic and protective functions.Self-reactive B cells achieve tolerance by receptor editing, a process that replaces V_H and/or V_L genes encoding the autoreactive receptor with genes that change or modify the self-reactivity (1). The process is stimulated by exposure to self-antigen (Ag) and is carried out by secondary rearrangement. The benefit of editing is that ongoing rearrangements can extinguish the autoreactive specificity. However, the immune system is neither perceptive nor perfect, and receptor editing also generates byproducts besides silenced anti-self receptors: The new receptors’ affinity for self may decrease below the threshold that triggers self tolerance but could convert to full-blown autoreactivity by somatic mutation in the periphery (2). Alternatively, editing can lead to rearrangements on a second allele, resulting in inclusion and the generation of bispecific, autoreactive B cells (2–6). In such allelically included cells, the autoreactive receptor is “diluted out” by the nonautoreactive one (6, 7), again resulting in the escape of an autoreactive B cell from self-tolerance.We are particularly interested in editing that leads to receptors with modified self activities. Certain combinations of anti-DNA V_H and editor V_L chains yield multireactive-autoreactive B-cell receptors (BCRs). However, despite the self-reactivity of their BCRs, these B cells escape further regulation and enter the periphery (8–10). The combination of V_H56R anti-DNA heavy (H) and Vκ38c editor light (L) chains is a case in point. This autoreactive Ab accumulates in the Golgi, presumably by binding to specific glycosaminoglycans expressed inside the secretory pathway (11). As a result, surface expression of the BCR is reduced, and B cells expressing V_H56R/Vκ38c escape from central tolerance.B cells with incompletely edited anti-DNA receptors are a ready source of potentially pathogenic Abs because arginines (R) in V_H function in an autonomous and additive manner as critical DNA binding residues (12). Thus, DNA binding is achieved without regard to most L chains (13). Most L chains sustain DNA binding when paired with V_H3H9 (14) or V_H56R (15). A few, however, can function as effective editors of anti-DNA reactivity (16). These anti-DNA editors are characterized by the presence of several aspartic acid (D) residues in their CDRs. The negatively charged Ds may block DNA binding by competing for the positive charges of Rs. The Vλx editor provides an example of just such an R–D interaction (Fig. 1; ref. 17). Vλx differs from other editor Vs by having Ds in CDR2 (L2) and in CDR3 (L3) (18). However, the high D content can be a liability: Vλx, presumably aided by the Ds, binds to cationic Ags such as MBP (Myelin Basic Protein) (19). Importantly, if the Ds and Rs do not complement each other, the Ab may be only partially or incompletely edited. The receptor of an incompletely edited B cell may still bind DNA through free R(s), MBP by free D(s) and a variety of other Ags.Open in a separate windowFig. 1.Example of interchain bonding between D and R. The crystal structure of MW1, an anti-polyQ Ab (17), reveals the interaction of D60 in the Vλx L chain and R96 in V_H. The side chains interact across the cleft separating V_H from V_L. We propose that an interaction between R and D side chains limits their availability for binding to Ag. In this way, editor L chains could reduce the propensity for DNA binding of R in anti-DNA H chains.Many anti-DNA are multireactive as R residues in the CDRs can also contribute to binding to anionic phospholipids such as cardiolipin and phosphatidylserine (PS). Indeed, the original 3H9 antibody was found to bind not only to dsDNA and chromatin, but also to cardiolipin (20). Site-directed mutagenesis experiments indicate that Rs in the H chain play a leading role in PS binding (21), whereas the L chain modulates affinity, similar to anti-DNA (8).The mechanism of editing anti-DNAs, and some antibodies to anionic lipids, appears to depend on strategically positioned L chain Ds, such as D96 in Vλx. Here, we exploit a natural example of structural heterogeneity in Vλx that alters L3 conformation and D96 position. The V-J rearrangement process generates Vλx junctions (Fig. 2) that are shorter by two codons or longer by two codons relative to the exact, canonical junction (CJ; ref. 22). This junctional diversity accounts, in part, for the binding differences between V_H56R/Vλx monoclonal Abs (23). Here, we compared Abs with the CJ of Vλx–Jλ2 to Abs with the two-codon insertion (+2) or deletion (−2). We studied the (self) Ag specificities of the three H/L pairs, and noted that the Abs differ in their binding to DNA, phospholipids and myosin. Of particular interest were differences in the binding of the Vλx variants to the Golgi and to glycosaminoglycans. We show that V_H56R/Vλx junctional variants that bind Golgi-associated Ags also accumulate inside cells, thereby recapitulating the result of V_H56R editing by the Vκ38c L chain (11). The intracellular sequestration may obviate the need for editing of these anti-DNA receptors, as they are diverted to Ags expressed in the Golgi. The common result of editing by the Vλx variants and Vκ38c is highly significant. The repeated observation that the two hybridomas accumulate intracellular IgM and release insoluble IgM complexes suggests that insoluble, amyloid-like IgM deposits arise as a biologically relevant and predictable outcome of anti-DNA receptor editing.Open in a separate windowFig. 2.Sequences of the Vλx–Jλ2 junctions characterized in this study. Editor Vλx L chains use the canonical junction (Top, CJ) or junctions that represent a deletion of two codons (Middle, −2) or insertion of two codons (Bottom, +2) relative to the CJ. The figure shows the partial nucleotide sequences of the Vλx and Jλ2 gene segments and the three rearrangements along with the amino acid sequences encoded by the rearrangements. The alternative junctions were generated by deletion of seven nucleotides upstream of the TAA codon of Vλx or by the insertion of two codons between Vλx and Jλ2, with likely involvement of P nucleotides. The TAT codon at the Vλx–Jλ2 junction is residue 104 if residues are sequentially counted starting with the amino terminus of Vλx. In a survey of Vλx junctions, 24 of 24 junctions that were analyzed were generated by joining thymidine from Jλ2 to the first adenine of the TAA terminator, thus generating a tyrosine (TAT) codon at position 104 and preventing premature termination (44).     

GSTM1 genotype is an independent prognostic factor in clear cell renal cell carcinoma

Purpose

Owing to dual functionality of cytosolic glutathione S-transferases (GSTs), they might affect both the development and the progression of renal cell carcinoma (RCC). However, the data on the prognostic value of GST polymorphism in patients with RCC are scarce. Hence, we evaluated the effect of GST gene variants on both the risk of RCC development and the postoperative prognosis in patients with clear cell RCC (ccRCC).