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Objective: To study the effects of natalizumab treatment on subgroups of circulating peripheral blood B cell populations., Methods: We studied the proportions and absolute numbers of CD19 + CD20 + , CD10 + , and CD5 + B cell populations, and determined very late activation antigen-4 and chemokine receptor CXCR3, CCR5, and CCR6 expression on B cells in the peripheral blood of 14 natalizumab-treated patients with relapsing-remitting multiple sclerosis. Five blood samples per patient were obtained longitudinally before and during the first year of treatment. Blood samples were analyzed by 6-color flow cytometry., Results: Proportions of B cells and CD10 + pre-B cells were significantly increased, and very late activation antigen-4 expression on the B cell surface was significantly decreased already after 1 week of natalizumab treatment. Natalizumab-induced sustained increase in the proportion and absolute number of CXCR3-expressing B cells was statistically significant after 1 month of treatment. There were no changes in the proportions of CCR5- or CCR6-expressing B cells., Conclusions: The rapid and persistent increase in circulating CXCR3-expressing B cells in response to natalizumab treatment possibly reflects the relevance of this chemokine receptor in controlling migration of B cells into the CNS in humans in vivo.

Multiparameter flow cytometry (MFC) and allele-specific oligonucleotide real-time quantitative PCR (ASO RQ-PCR) are the two most sensitive methods to detect minimal residual disease (MRD) in multiple myeloma (MM). We compared these methods in 129 paired post-therapy samples from 22 unselected, consecutive MM patients in complete/near complete remission. Appropriate immunophenotypic and ASO RQ-PCR-MRD targets could be detected and MRD analyses constructed for all patients. The high PCR coverage could be achieved by gradual widening of the primer sets used for clonality detection. In addition, for 13 (55%) of the patients, reverse orientation of the ASO primer and individual design of the TaqMan probe improved the sensitivity and specificity of ASO RQ-PCR analysis. A significant nonlinear correlation prevailed between MFC-MRD and PCR-MRD when both were positive. Discordance between the methods was found in 32 (35%) paired samples, which were negative by MFC-MRD, but positive by ASO RQ-PCR. The findings suggest that with the described technique, ASO RQ-PCR can be constructed for all patients with MM. ASO RQ-PCR is slightly more sensitive in MRD detection than 6-10-color flow cytometry. Owing to technical demands ASO RQ-PCR could be reserved for patients in immunophenotypic remission, especially in efficacy comparisons between different drugs and treatment modalities.

Objective: To evaluate [1] the effects of levels of sex hormone-binding globulin (SHBG), albumin, and total testosterone on the distribution of testosterone between SHBG-bound and non-SHBG-bound fractions; [2] the independent effects of polycystic ovary syndrome (PCOS) and body mass index on serum levels of total testosterone, non-SHBG-bound testosterone, SHBG, and albumin; and [3] the usefulness of levels of total testosterone and non-SHBG-bound testosterone and of the free androgen index in the diagnosis of PCOS., Design: Retrospective clinical study., Setting: An academic research environment., Patient(s): Forty-three women with oligomenorrhea and PCOS. Twenty-five women with regular menstrual cycles and without hirsutism served as controls., Intervention(s): None., Main Outcome Measure(s): Levels of non-SHBG-bound testosterone, total testosterone, SHBG, and albumin in serum., Result(s): Levels of total testosterone and non-SHBG-bound testosterone, and the free androgen index were higher in patients with PCOS than in healthy controls. PCOS did not have an effect on the levels of SHBG or albumin, or on the percentage of non-SHBG-bound testosterone. Levels of SHBG and albumin were inversely related to body mass index. The percentage and concentration of non-SHBG-bound testosterone and the free androgen index were directly related to body mass index. Hirsutism did not have an effect on any outcome measure., Conclusion(s): The distribution of total testosterone into SHBG-bound and non-SHBG-bound fractions is associated with body mass index, not with PCOS. The high levels of non-SHBG-bound testosterone and the high free androgen index in patients with PCOS reflect mainly high levels of total testosterone. Thus, the measurement of levels of non-SHBG-bound testosterone and the calculation of the free androgen index provide no further information in the diagnosis of PCOS beyond that provided by the measurement of levels of total testosterone.

The temporal and spatial expression of thirteen novel spermatid-specific genes corresponding to cDNA clones isolated from an adult mouse testis library was analyzed. Northern analysis of RNA from seminiferous tubules at defined stages of the rat and mouse seminiferous epithelial cycle and in situ hybridization of testis sections revealed that these mRNAs were expressed in a stage-specific manner. The expression of all mRNAs was first detected in early round spermatids, and it increased to abundance during stages VII-VIII of the epithelial cycle. Twelve out of thirteen mRNAs were found not only in round spermatids but also in transcriptionally inactive elongated spermatids, suggesting that they are stored and regulated at the translational level. The variation in the length of the poly(A) tail was detected for four of the transcripts, represented by cDNA clones MTEST70, MTEST627, MTEST641, and MTEST643 at defined stages of the cycle. Similarity in the stage-specific expression pattern displayed by this group of haploid-specific genes strongly suggests the presence of common regulatory mechanisms that act during spermiogenesis, and these genes also provide a means for further studies of these mechanisms.

In this study we investigated the expression of the immediate early genes (IEGs) c-fos, c-jun, and junD on mRNA and protein levels during the spermatogenic cycle of the rat using Northern blotting, in situ hybridization, and immunocytochemistry. The expression of these IEG mRNAs and proteins exhibited stage-specific variations. The results suggest that IEGs take part in transcriptional events that are involved in regulating the proliferation and differentiation of spermatogenic cells during specific stages of the cycle of the seminiferous epithelium.

Mad is a basic region helix-loop-helix leucine zipper transcription factor which can dimerize with the Max protein and antagonize transcriptional activation by the Myc-Max transcription factor heterodimer. While the expression of Myc is necessary for cell proliferation, the expression of Mad is induced upon differentiation of at least some leukemia cell lines. Here, the expression of the mad gene has been explored in developing mouse tissues. During organogenesis in mouse embryos mad mRNA was predominantly expressed in the liver and in the mantle layer of the developing brain. At later stages mad expression was detected in neuroretina, epidermis, and whisker follicles, and in adult mice mad was expressed at variable levels in most organs analyzed. Interestingly, in the skin mad was highly expressed in the differentiating epidermal keratinocytes, but not in the underlying proliferating basal keratinocyte layer. Also, in the gut mad mRNA was abundant in the intestinal villi, where cells cease proliferation and differentiate, but not in the crypts, where the intestinal epithelial cells proliferate. In the testis, mad expression was associated with the completion of meiosis and early development of haploid cells. In cell culture, Mad inhibited colony formation of a mouse keratinocyte cell line and rat embryo fibroblast transformation by Myc and Ras. The pattern of mad expression in tissues and its ability to inhibit cell growth in vitro suggests that Mad can cause the cessation of cell proliferation associated with cell differentiation in vivo.