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Benign melanocytic nevi are slowly growing acquiredor congenital tumors with varied morphology,commonly encountered in dermatology clinics. Anytumor with rapid clinical growth must be assessedcarefully in order to exclude malignancy. We report awoman with a histopathologically benign intradermalnevus that presented as a rapidly evolving largecutaneous mass on the ear. Owing to the discrepancybetween the clinical and histopathological findings,an extensive histopathological work-up involvingmany deeper sections, immunohistochemical stains,and fluorescent in situ hybridization (FISH) analysiswas conducted in order to rule out malignancy.

Treatment of cancer in children is increasingly successful but leaves many prepubertal boys suffering from infertility or subfertility later in life. A current strategy to preserve fertility in these boys is to cryopreserve a testicular biopsy prior to treatment with the expectation of future technologies allowing for the reintroduction of stem cells and restoration of spermatogenesis. Spermatogonial stem cells (SSCs) form the basis of male reproduction, differentiating into all germ cell types, including mature spermatozoa and can regenerate spermatogenesis following transplantation into an infertile testis. Here, we demonstrate that rat SSCs frozen for more than 20 years can be transplanted into recipient mice and produce all differentiating germ cell types. However, compared with freshly isolated cells or those frozen for a short period of time, long-frozen cells do not colonize efficiently and showed reduced production of spermatids. Single-cell RNA sequencing revealed similar profiles of gene expression changes between short- and long-frozen cells as compared with fresh immediately after thawing. Conversely, following transplantation, long-frozen samples showed enhanced stem cell signaling in the undifferentiated spermatogonia compartment, consistent with self-renewal and a lack of differentiation. In addition, long-frozen samples showed fewer round spermatids with detectable protamine expression, suggesting a partial block of spermatogenesis after meiosis resulting in a lack of elongating spermatids. These findings strongly suggest that prolonged cryopreservation can impact the success of transplantation to produce spermatogenesis, which may not be revealed by analysis of the cells immediately after thawing. Our analysis uncovered persistent effects of long-term freezing not found in other cryopreservation studies that lacked functional regeneration of the tissue and this phenomenon must be accounted for any future therapeutic application. This study shows that spermatogonial stem cells frozen for more than 20 years can repopulate the niche when transplanted into an infertile host, but with lower efficiency and displaying abnormal spermatogenesis compared to fresh or short-term frozen cells. Single-cell RNA sequencing reveals details of the dysregulated signaling.

With improved outcomes associated with radiotherapy (RT), post-irradiation tumors are increasingly seen in long-term cancer survivors. We report a case of a young woman who presented with a three-year history of a vascular lesion on the temple, previously irradiated for a childhood brain tumor. The history of radiation, the clinical appearance, and the biopsy findings of an atypical vascular proliferation in the dermis, were worrisome for a malignant vascular neoplasm and prompted surgical excision. However, further tissue analysis of the excised specimen confirmed a benign atypical vascular lesion (AVL) overlying a banal pilar cyst. Distinguishing post-radiation benign from malignant vascular lesions can be difficult because they share overlapping clinical and histopathologic features. Thus, any vascular lesion that occurs in a previously irradiated field should be excised completely with tumor-free margins and examined histologically.

The chemokine (C-X-C) receptor type 4 (CXCR4) is an early marker of primordial germ cells (PGCs) essential for their migration and colonization of the gonads. In spermatogonial stem cells (SSCs), the expression of CXCR4 is promoted by the self-renewal factor, glial cell line-derived neurotrophic factor (GDNF). Here, we demonstrate an important role of CXCR4 during donor mouse SSCs reoccupation of the endogenous niche in recipient testis. Silencing of CXCR4 expression in mouse SSCs dramatically reduced the number of donor stem cell-derived colonies, whereas colony morphology and spermatogenesis were comparable to controls. Inhibition of CXCR4 signaling using a small molecule inhibitor (AMD3100) during the critical window of homing also significantly lowered the efficiency of donor-derived SSCs to establish spermatogenic colonies in recipient mice; however, the self-renewal of SSCs was not affected by exposure to AMD3100. Rather, in vitro migration assays demonstrate the influence of CXCR4-CXCL12 signaling in promoting germ cell migration. Together, these studies suggest that CXCR4-CXCL12 signaling functions to promote homing of SSCs towards the stem cell niche and plays a critical role in reestablishing spermatogenesis.

The dual-function Rel(sub Mtb) protein from Mycobacterium tuberculosis catalyzes both the synthesis and hydrolysis of (p)ppGpp, the effector of the stringent response. Purified and characterized fragment of the 738-amino acid Rel(sub Mtb) protein is presented and it is confirmed that the hypothesis that amino acid fragment 1-394 contains both synthesis and hydrolysis activities.

Opposing Rel(sub.Mtb) activities by the aminoacylation state of a tRNA.ribosome.RNAm Rel(sub.Mtb) complex and their differential regulation in Mycobacterium tuberculosis are discussed. The differential Rel(sub.Mtb) control gives the Mtb ribosomal complex a new regulatory role for cellular metabolism responding to growth conditions that may be present in the dormant Mtb lesion.

Spermatogonial stem cells (SSCs) are at the foundation of the highly productive spermatogenic process that continuously produces male gametes throughout postnatal life. However, experimental evaluation of SSCs in postnatal testes is complicated because these cells are extremely rare and few defining morphology or biochemical characteristics are known. In this study, we used the spermatogonial transplantation functional assay, combined with fluorescence-activated cell sorting (FACS) analysis to identify cellular, biochemical and surface antigenic characteristics of SSCs in rat testes during development. Our results demonstrated that forward scatter [(FSc).sup.hi], side scatter [(SSc).sup.hi], mitochondria membrane potential [([DELTA][PSI]m).sup.1o], Ep-CAM+, Thy-1+, [beta]3-integrin+ stem cells in neonate rat testes become [SSc.sup.1o], [DELTA][PSI][h.sup.hi], Ep-CAM+, Thy-[1.sup.1o], [beta]3-integrin- stem cells in pup rat testes. Furthermore, prospective identification of rat testis cell populations (Ep-CAM+), highly enriched for SSCs (1 in 13 for neonate; 1 in 8.5 for pup) enabled us to predict the Thy-1 and [beta]3-integrin status of stem cells in neonate and pup testes, which was subsequently confirmed by transplantation analyses. Systematic characterization of SSCs enabled the production of testis cell populations highly enriched (up to 120-fold) for SSCs and will facilitate future investigations of functional and genomic characteristics. Keywords: Stem cell; Spermatogonial stem cell; Spermatogenesis; Germline; Development; Rat; Transplantation; Testis; Enrichment

Adult tissue stem cells self-renew and differentiate in a way that exactly meets the biological demand of the dependent tissue. We evaluated spermatogonial stem cell (SSC) activity in the developing rat testis and the quality and accessibility of the stem cell niche in wild type, and two busulfan-treated models of rat pup recipient testes using an SSC transplantation technique as a functional assay. While our results revealed a 69-fold increase in stem cell activity during rat testis development from neonate to adult, only moderate changes in SSC concentration were observed, and stem cells from neonate, pup, and adult donor testes produce spermatogenic colonies of similar size. Analysis of the stem cell niche in recipient rat testes demonstrated that pup testes support high levels of donor stem cell engraftment when endogenous germ cells are removed or compromised by busulfan treatment. Fertility was established when rat pup donor testis cells were transplanted into fetal- or pup-busulfan-treated recipient rat pup testes, and the donor genotype was transmitted to subsequent generations. These results provide insight into stem cell/niche interactions in the rat testis and demonstrate that techniques originally developed in mice can be extended to other species for regenerative medicine and germline modification. Keywords: Stem cell; Spermatogonial stem cell; Niche; Spermatogenesis; Development; Rat; Fertility; Transplantation; Testis

Long-term survival of nonreplicating Mycobacterium tuberculosis (Mtb) is ensured by the coordinated shutdown of active metabolism through a broad transcriptional program called the stringent response. In Mtb, this response is initiated by the enzymatic action of Re[l.sub.Mtb] and deletion of re[l.sub.Mtb] produces a strain (H37Rv1[DELTA]re[l.sub.Mtb]) severely compromised in the maintenance of long-term viability. Although aerosol inoculation of mice with H37Rv[DELTA]re[l.sub.Mtb] results in normal initial bacterial growth and containment, the ability of this strain to sustain chronic infection is severely impaired. Significant histopathologic differences were noted in lungs and spleens of mice infected with H37Rv[DELTA]re[l.sub.Mtb] compared with controls throughout the course of the infection. Microarray analysis revealed that H37Rv[DELTA]re[l.sub.Mtb] suffers from a generalized alteration of the transcriptional apparatus, as well as specific changes in the expression of virulence factors, cell-wall biosynthetic enzymes, heat shock proteins, and secreted antigens that may alter immune recognition of the recombinant organism. Thus, Re[l.sub.Mtb] is critical for the successful establishment of persistent infection in mice by altering the expression of antigenic and enzymatic factors that may contribute to successful latent infection.

Research describes cloning of the cydA and cydB genes that code for the two subunits of the cytochorme bd oxidase in Mycobacterium smegmatis along with the analysis of their expression and phenotypic characterization of the mutants. Data indicate that cytochrome bd oxidase has a role in the respiration of the bacterium.

Results indicate that Mycobacterium tuberculosis adaptation to long-term survival in asymptomatic patients is mediated by hyperphosphorylated guanine controlling gene expresson. Data further point out that rel(sub)Mtb protein generates the hyperphosphorylated guanine species under nutrient starvation conditions.

Author(s): Takehiko Ogawa [1, 2]; Ina Dobrinski [1, 3]; Mary R. Avarbock [1]; Ralph L. Brinster (corresponding author) [1] Approximately 50% of human infertility is attributable to male defects, 70-90% [...]

Although spermatogenesis is essential for reproduction, little is known about spermatogonial stem cells. These cells provide the basis for spermatogenesis throughout adult life by undergoing self-renewal and by providing progeny that differentiate into spermatozoa. A major impediment to our understanding of the biology of these stem cells is the inability to distinguish them from spermatogonia that are committed to differentiation. We made use of the known association of stem cells with basement membranes and our spermatogonial transplantation assay system to identify specific molecular markers on the stem cell surface. Selection of mouse testis cells with anti-[[Beta].sub.1]- or anti-[[Alpha].sub.6]-integrin antibody, but not anti-c-kit antibody, produced cell populations with a significantly enhanced ability to colonize recipient testes and generate donor cell-derived spermatogenesis. We demonstrate spermatogonial stem cell-associated antigens by using an assay system based on biological function. Furthermore, the presence of surface integrins on spermatogonial stem cells suggests that these cells share elements of a common molecular machinery with stem cells in other tissues.

A research study was conducted to further characterize the genes, nrdF1 and nrdF2 which are encoding a small unit (R2) of ribonucleotide reductase (RR) from the Mycobacterium tuberculosis (M tuberculosis). Some of the methods used were the isolation of partial R2 sequences, Southern analysis, PCR analysis of M tuberculosis strains, expression of M tuberculosis R2-1 and R2-2, purification of recombinant M tuberculosis R2-2 and peptide purification. Results showed that M tuberculosis nrdEF system seems to encode the active species of RR.

Rat spermatogenesis in mouse testis is possible by transplanting testis cells from transgenic rats to the testes of immunodeficient mice. The rat spermatozoa produced in mouse testis are normal, even though rat spermatogenesis requires almost 50% more time than that of mouse. Observations show that mouse Sertoli cells support rat spermatogenesis. The male germ line stem cells could find applications in agriculture, biology and medicine due to their ability to replicate throughout their life.

A study of stem cell spermatogenesis through the transplantation of these cells from donor testis into infertile recipient testis reveals that spermatogonia repopulate the recipient testis more efficiently than gonocytes and that the transplanted donor spermatogonia produce daughter cells. Spermatogenesis occurs through colonization of the seminiferous tubules of the recipient testis by the transplanted stem cell spermatogonia, the colonization efficiency being low for cells from new-born mice. Nearly 80% of the daughter cells of recipients that undergo transplantation and exhibit endogenous spermatogenesis arise from donor-derived spermatogonia.