Early Outpatient Treatment for Covid-19 with Convalescent Plasma.

BACKGROUND: Polyclonal convalescent plasma may be obtained from donors who have recovered from coronavirus disease 2019 (Covid-19). The efficacy of this plasma in preventing serious complications in outpatients with recent-onset Covid-19 is uncertain. METHODS: In this multicenter, double-blind, randomized, controlled trial, we evaluated the efficacy and safety of Covid-19 convalescent plasma, as compared with control plasma, in symptomatic adults (≥18 years of age) who had tested positive for severe acute respiratory syndrome coronavirus 2, regardless of their risk factors for disease progression or vaccination status. Participants were enrolled within 8 days after symptom onset and received a transfusion within 1 day after randomization. The primary outcome was Covid-19-related hospitalization within 28 days after transfusion. RESULTS: Participants were enrolled from June 3, 2020, through October 1, 2021. A total of 1225 participants underwent randomization, and 1181 received a transfusion. In the prespecified modified intention-to-treat analysis that included only participants who received a transfusion, the primary outcome occurred in 17 of 592 participants (2.9%) who received convalescent plasma and 37 of 589 participants (6.3%) who received control plasma (absolute risk reduction, 3.4 percentage points; 95% confidence interval, 1.0 to 5.8; P = 0.005), which corresponded to a relative risk reduction of 54%. Evidence of efficacy in vaccinated participants cannot be inferred from these data because 53 of the 54 participants with Covid-19 who were hospitalized were unvaccinated and 1 participant was partially vaccinated. A total of 16 grade 3 or 4 adverse events (7 in the convalescent-plasma group and 9 in the control-plasma group) occurred in participants who were not hospitalized. CONCLUSIONS: In participants with Covid-19, most of whom were unvaccinated, the administration of convalescent plasma within 9 days after the onset of symptoms reduced the risk of disease progression leading to hospitalization. (Funded by the Department of Defense and others; CSSC-004 ClinicalTrials.gov number, NCT04373460.).

Natural history of clonal haematopoiesis seen in real-world haematology settings.

Recursive partitioning of healthy consortia led to the development of the Clonal Hematopoiesis Risk Score (CHRS) for clonal haematopoiesis (CH); however, in the practical setting, most cases of CH are diagnosed after patients present with cytopenias or related symptoms. To address this real-world population, we characterize the clinical trajectories of 94 patients with CH and distinguish CH harbouring canonical DNMT3A/TET2/ASXL1 mutations alone ('sole DTA') versus all other groups ('non-sole DTA'). TET2, rather than DNMT3A, was the most prevalent mutation in the real-world setting. Sole DTA patients did not progress to myeloid neoplasm (MN) in the absence of acquisition of other mutations. Contrastingly, 14 (20.1%) of 67 non-sole DTA patients progressed to MN. CHRS assessment showed a higher frequency of high-risk CH in non-sole DTA (vs. sole DTA) patients and in progressors (vs. non-progressors). RUNX1 mutation conferred the strongest risk for progression to MN (odds ratio [OR] 10.27, 95% CI 2.00-52.69, p = 0.0053). The mean variant allele frequency across all genes was higher in progressors than in non-progressors (36.9% ± 4.62% vs. 24.1% ± 1.67%, p = 0.0064). This analysis in the post-CHRS era underscores the natural history of CH, providing insight into patterns of progression to MN.

Symptom Duration and Resolution With Early Outpatient Treatment of Convalescent Plasma for Coronavirus Disease 2019: A Randomized Trial.

BACKGROUND: Coronavirus disease 2019 (COVID-19) convalescent plasma (CCP) reduces hospitalizations among outpatients treated early after symptom onset. It is unknown whether CCP reduces time to symptom resolution among outpatients. METHODS: We evaluated symptom resolution at day 14 by trial arm using an adjusted subdistribution hazard model, with hospitalization as a competing risk. We also assessed the prevalence of symptom clusters at day 14 between treatments. Clusters were defined based on biologic clustering, impact on ability to work, and an algorithm. RESULTS: Among 1070 outpatients followed up after transfusion, 381 of 538 (70.8%) receiving CCP and 381 of 532 (71.6%) receiving control plasma were still symptomatic (P = .78) at day 14. Associations between CCP and symptom resolution by day 14 did not differ significantly from those in controls after adjustment for baseline characteristics (adjusted subdistribution hazard ratio, 0.99; P = .62). The most common cluster consisted of cough, fatigue, shortness of breath, and headache and was found in 308 (57.2%) and 325 (61.1%) of CCP and control plasma recipients, respectively (P = .16). CONCLUSIONS: In this trial of outpatients with early COVID-19, CCP was not associated with faster resolution of symptoms compared with control. Overall, there were no differences by treatment in the prevalence of each symptom or symptom clusters at day 14. CLINICAL TRIALS REGISTRATION: NCT04373460.

Transfusing Convalescent Plasma as Post-Exposure Prophylaxis Against Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Infection: A Double-Blinded, Phase 2 Randomized, Controlled Trial.

BACKGROUND: The efficacy of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) convalescent plasma (CCP) for preventing infection in exposed, uninfected individuals is unknown. CCP might prevent infection when administered before symptoms or laboratory evidence of infection. METHODS: This double-blinded, phase 2 randomized, controlled trial (RCT) compared the efficacy and safety of prophylactic high titer (≥1:320 by Euroimmun ELISA) CCP with standard plasma. Asymptomatic participants aged ≥18 years with close contact exposure to a person with confirmed coronavirus disease 2019 (COVID-19) in the previous 120 hours and negative SARS-CoV-2 test within 24 hours before transfusion were eligible. The primary outcome was new SARS-CoV-2 infection. RESULTS: In total, 180 participants were enrolled; 87 were assigned to CCP and 93 to control plasma, and 170 transfused at 19 sites across the United States from June 2020 to March 2021. Two were excluded for screening SARS-CoV-2 reverse transcription polymerase chain reaction (RT-PCR) positivity. Of the remaining 168 participants, 12/81 (14.8%) CCP and 13/87 (14.9%) control recipients developed SARS-CoV-2 infection; 6 (7.4%) CCP and 7 (8%) control recipients developed COVID-19 (infection with symptoms). There were no COVID-19-related hospitalizations in CCP and 2 in control recipients. Efficacy by restricted mean infection free time (RMIFT) by 28 days for all SARS-CoV-2 infections (25.3 vs 25.2 days; P = .49) and COVID-19 (26.3 vs 25.9 days; P = .35) was similar for both groups. CONCLUSIONS: Administration of high-titer CCP as post-exposure prophylaxis, although appearing safe, did not prevent SARS-CoV-2 infection. CLINICAL TRIALS REGISTRATION: NCT04323800.

How do I implement an outpatient program for the administration of convalescent plasma for COVID-19?

Convalescent plasma, collected from donors who have recovered from a pathogen of interest, has been used to treat infectious diseases, particularly in times of outbreak, when alternative therapies were unavailable. The COVID-19 pandemic revived interest in the use of convalescent plasma. Large observational studies and clinical trials that were executed during the pandemic provided insight into how to use convalescent plasma, whereby high levels of antibodies against the pathogen of interest and administration early within the time course of the disease are critical for optimal therapeutic effect. Several studies have shown outpatient administration of COVID-19 convalescent plasma (CCP) to be both safe and effective, preventing clinical progression in patients when administered within the first week of COVID-19. The United States Food and Drug Administration expanded its emergency use authorization (EUA) to allow for the administration of CCP in an outpatient setting in December 2021, at least for immunocompromised patients or those on immunosuppressive therapy. Outpatient transfusion of CCP and infusion of monoclonal antibody therapies for a highly transmissible infectious disease introduces nuanced challenges related to infection prevention. Drawing on our experiences with the clinical and research use of CCP, we describe the logistical considerations and workflow spanning procurement of qualified products, infrastructure, staffing, transfusion, and associated management of adverse events. The purpose of this description is to facilitate the efforts of others intent on establishing outpatient transfusion programs for CCP and other antibody-based therapies.

Robinow syndrome: Genital analysis, genetic heterogeneity, and associated psychological impact.

Robinow syndrome (RS) is a rare, pleiotropic genetic disorder. While it has been reported that males with Robinow syndrome may have genitourinary atypicalities, these have not been systematically studied. We hypothesized that the underlying gene involved plays a role in the clinical variability of associated genital findings and that the phenotypic appearance of the genitalia in RS may have a psychological impact. Urologic-specific examination consisted of detailed examination and a questionnaire to investigate the psychological impact of the genital phenotype. Nine males agreed to a full evaluation. Average age was 19.9 years, penile length was 32.5 mm, stretched length 53 mm, and width 24.4 mm. Penile transposition occurred in all 9 male who allowed full examination. Undescended testicles were noted in 4/10, testicular atrophy in 5/9, buried penis in 7/9, hypospadias in 5/8, and a large penopubic gap (space between dorsum of penis base and pubic bone) in 5/6. In this cohort, 78% answered our semi-quantitative pilot questionnaire that identified diminished sexuality, sexual function, and self-perception. In conclusion, RS has unique, hallmark genital findings including penile transposition, buried penis, undescended testes, and large penopubic gaps. Genital phenotype in males was not shown to correlate with the specific gene involved for each patient. Surgical approaches and other interventions should be studied to address the findings of decreased sexuality and self-perception. It is the authors' opinion that intervention to provide the appearance of penile lengthening be postponed until puberty to allow for maximal natural phallic growth.

Prevalence of high-risk bladder categorization with prenatal and postnatal myelomeningocele repair types.

INTRODUCTION: Urologic substudies of prenatal myelomeningocele (MMC) closure have focused primarily on continence without significant clinical benefit. Fetoscopic MMC repair (FMR) is a newer form of prenatal intervention and touts added benefits to the mother, but urological outcomes have yet to be analyzed. We set out to focus on bladder safety rather than continence and examined bladder outcomes with different prenatal MMC repairs (FMR and prenatal open [POMR]) and compared bladder-risk-categorization to traditional postnatal repair (PSTNR). METHODS: An IRB-approved retrospective analysis of all patients undergoing all forms of MMC repairs with inclusion and exclusion criteria based on the MOMS trial was performed. Bladder safety assessment required initial urodynamic studies (UDS), renal bladder ultrasound (RBUS), and/or voiding cystourethrogram (VCUG) within the 1st year of life. Follow-up analyses within the cohorts required follow-up studies within 18 months after initial evaluations. Outcomes assessed included bladder-risk-categorization based on the CDC UMPIRE study (high, intermediate, and safe), hydronephrosis (HN), and vesicoureteral reflux (VUR). A single reader evaluated each UDS. RESULTS: Initial UDS in 93 patients showed that the prevalence of high-risk bladders were 35% FMR versus 36% PSTNR and 60% POMR. Follow-up UDS showed only 8% of FMR were high-risk compared to 35% POMR and 36% PSTNR. Change from initial to follow-up bladder-risk-category did not reach significance (p = .0659); however, 10% PSTNR worsened to high-risk on follow-up, compared to none in either prenatal group. Subanalysis of follow-up UDS between the prenatal cohorts also was not significant (p = .055). Only 8% of FMR worsened or stayed high-risk compared to 35% with POMR (p = .1). HN was significantly different at initial and subsequent follow up between the groups with the least in the FMR group. CONCLUSIONS: Early outcome UDS analyses demonstrated lower incidence of high-risk bladders in FMR patients with a trend toward clinically significant improvement compared to POMR in regard to all evaluated metrics. Larger, prospective, confirmatory studies are needed to further evaluate the potential benefits on FMR on bladder safety and health.

Analysing online Twitter discussions of bedwetting via a condition-specific hashtag (#Bedwetting).

AIM: Bedwetting is a common paediatric condition. #Bedwetting has been established as the official hashtag to structure Twitter discussions about the condition. We analysed online Twitter discussions for #Bedwetting. METHODS: Symplur, a Twitter analytics service was employed to aggregate Twitter activity, users and content including #Bedwetting, between October 2013 and November 2018. Activity was analysed via tweet volume and user adoption. Users were assorted using geographic location, occupation and affiliation data. Content in #Bedwetting Tweets was undertaken by retrieving information about retweets, links, frequently used words and hashtags. RESULTS: A total of 101 412 tweets and 9957 users utilising #Bedwetting were identified. Most tweets were sent with links (93%). The average ± SD number of tweets using #Bedwetting per month increased from 96 ± 87 in 2013 to 2935 ± 1644 in 2015. Tweet volume decreased to 1960 ± 257 in 2016 and subsequently increased to 2901 ± 1110 in 2017. New users increased from 4 in 2013 to 9957 users in 2018. Users tweeted from 69 countries. Advocacy organisations comprised 35% of the top 100 influencers. Common words in #Bedwetting tweets were 'potty', 'best' and 'training'. Popular associated hashtags were #Pottytraining, #Solutions and #Moms. Hyperlinks in #Bedwetting tweets included advocacy, academic and commercial websites. CONCLUSIONS: Our analysis of #Bedwetting highlights that Twitter is frequently used to discuss the condition's diagnosis and management. Various stakeholders in health care are utilising the platform to build awareness about bedwetting. We identified that Twitter is being employed to drive web traffic to other internet websites.

Elotuzumab-based maintenance therapy following autologous stem cell transplant in multiple myeloma deepens post-transplant responses.

BACKGROUND: Post-transplant maintenance provides progression-free survival benefit in multiple myeloma (MM). Here we report our institution's experience with elotuzumab-based maintenance following autologous stem cell transplant. METHODS: We retrospectively evaluated the outcomes of MM patients who were started on elotuzumab-based maintenance (elotuzumab/lenalidomide/dexamethasone, elotuzumab/bortezomib/dexamethasone, or elotuzumab/bortezomib/methylprednisolone) following transplant (N = 7). Baseline characteristics, treatment response, survival, and adverse events were reviewed. RESULTS: Median age was 68 (56-81) years at the time of transplant, and median lines of induction therapy was 2 (1-6). Three patients (42.9%) had high-risk cytogenetics and five (71.4%) had stage II or greater disease at diagnosis. At a median follow-up of 24 months (12-50), five patients (71.4%) had improvement of quality of response, with a combined CR or VGPR rate increasing from 57.1% to 100% (CR = 3, VGPR = 4). All patients were alive without relapse or progression at the time of this analysis. Grade 3-4 adverse events were observed in three (42.9%) patients. None of the patients discontinued the treatment due to intolerance. CONCLUSIONS: Our study demonstrates that elotuzumab-based maintenance may deepen response post-transplant in MM and can be safely administered even in older patients. Given its unique action and rare side effects, further studies of elotuzumab in the post-transplant setting are warranted.

Establishment and functional characterization of a murine primary Sertoli cell line deficient of connexin43.

The Sertoli cell (SC) specific connexin43 (Cx43) knockout (SCCx43KO) mouse line is ideal to gain insight into the mechanistic gap junction formation in SC and the seminiferous epithelium. A method for developing primary SC cultures from these mice was established, validated and successfully characterized via polymerase chain reaction, immunohistochemistry, immunofluorescence (IF), and Western blots (WB). It was evident that both knockout (KO) and wild-type (WT) primary cell cultures were similar in morphology. These highly pure SC cultures were subjected to cell proliferation assays indicating no notable proliferation in cultures of both genotypes. Measurements of cell monolayer integrity indicated significant increases in transepithelial electrical resistance and consequently in tight junction expression of the KO cultures. Using semi-quantitative WB and IF, tight junction protein claudin-11 was analyzed. These results support a role for Cx43 in regulating blood-testis barrier (BTB) function, composition, and dynamics in vitro. Thus, the SC deficient Cx43 cell cultures may provide a valuable in vitro tool for a better understanding of the mechanistic role of Cx43 in spermatogenesis and BTB assembly.

Robot-assisted laparoscopic ureteral reimplantation in children: a valuable alternative to open surgery.

INTRODUCTION: Robot-assisted laparoscopic surgical systems have led to new minimally invasive options for complex reconstructive procedures in children including for vesicoureteral reflux (VUR). Robot-assisted laparoscopic ureteral reimplantation has been shown to be a viable minimally invasive surgical option for children with VUR. However, higher-than-expected complication rates and sub-optimal reflux resolution rates at some centers have also been reported. METHODS: This article provides a focused literature review as well as current perspectives on open reimplantation and robot-assisted laparoscopic ureteral reimplantation as non-endoscopic surgical options for pediatric VUR. RESULTS: The heterogeneity of surgical outcomes may, in part, be due to the learning curve inherent with all new technology and procedures. As a result, the current gold standard surgical option for VUR continues to be open ureteral reimplantation. While it remains to be seen if robot-assisted laparoscopic surgery will gradually replace open surgery as the most utilized surgical option for VUR in pediatric patients, robot-assisted laparoscopic ureteral reimplantation with the current robotic surgical systems may be just one step toward an eventual minimally invasive option that all experienced surgeons can offer with the requisite high success rates and low major complication rates. CONCLUSION: Robot-assisted laparoscopic ureteral reimplantation remains a viable minimally invasive surgical option for children with VUR, but with the expected learning curve associated with all new technologies.

Effect of CYP3A4, CYP3A5, and ABCB1 Polymorphisms on Intravenous Tacrolimus Exposure and Adverse Events in Adult Allogeneic Stem Cell Transplant Patients.

Pharmacogenetics influences oral tacrolimus exposure; however, little data exist regarding i.v. tacrolimus. We investigated the impact of genetic polymorphisms in CYP3A4, CYP3A5, and ABCB1 on i.v. tacrolimus exposure and toxicity in adult patients receiving an allogeneic hematopoietic stem cell transplant for hematologic malignancies. Germline DNA was extracted from buccal swabs and genotyped for CYP3A4, CYP3A5, and ABCB1 polymorphisms. Continuous i.v. infusion of tacrolimus .03 mg/kg/day was initiated on day +5 post-transplant, and steady-state blood concentrations were measured 4days later. We evaluated the association between phenotypes and prevalence of nontherapeutic target concentrations (below or above 5 to 15 ng/mL) as well as tacrolimus-related toxicities. Of 63 patients, 28.6% achieved the target concentration; 71.4% were >15ng/mL, which was more common in CYP3A4 intermediate/normal metabolizers (compared with rapid) and those with at least 1 ABCB1 C2677T loss-of-function allele (P < .05). ABCB1 C2677T was significantly associated with concentrations >15ng/mL (odds ratio, 6.2; 95% confidence interval, 1.8 to 23.6; P = .004) and tacrolimus-related toxicities (odds ratio, 7.5; 95% confidence interval, 1.6 to 55.2; P = .02). ABCB1 C2677T and CYP3A4 are important determinants of i.v. tacrolimus exposure, whereas ABCB1 C2677T also impacts tacrolimus-related toxicities in stem cell transplants.

Blood-testis barrier and Sertoli cell function: lessons from SCCx43KO mice.

The gap junction protein connexin43 (CX43) plays a vital role in mammalian spermatogenesis by allowing for direct cytoplasmic communication between neighbouring testicular cells. In addition, different publications suggest that CX43 in Sertoli cells (SC) might be important for blood-testis barrier (BTB) formation and BTB homeostasis. Thus, through the use of the Cre-LoxP recombination system, a transgenic mouse line was developed in which only SC are deficient of the gap junction protein, alpha 1 (Gja1) gene. Gja1 codes for the protein CX43. This transgenic mouse line has been commonly defined as the SC specific CX43 knockout (SCCx43KO) mouse line. Within the seminiferous tubule, SC aid in spermatogenesis by nurturing germ cells and help them to proliferate and mature. Owing to the absence of CX43 within the SC, homozygous KO mice are infertile, have reduced testis size, and mainly exhibit spermatogenesis arrest at the level of spermatogonia, seminiferous tubules containing only SC (SC-only syndrome) and intratubular SC-clusters. Although the SC specific KO of CX43 does not seem to have an adverse effect on BTB integrity, CX43 influences BTB composition as the expression pattern of different BTB proteins (like OCCLUDIN, ß-CATENIN, N-CADHERIN, and CLAUDIN11) is altered in mutant males. The supposed roles of CX43 in dynamic BTB regulation, BTB assembly and/or disassembly and its possible interaction with other junctional proteins composing this unique barrier are discussed. Data collectively indicate that CX43 might represent an important regulator of dynamic BTB formation, composition and function.

Association of acute myeloid leukemia's most immature phenotype with risk groups and outcomes.

The precise phenotype and biology of acute myeloid leukemia stem cells remain controversial, in part because the "gold standard" immunodeficient mouse engraftment assay fails in a significant fraction of patients and identifies multiple cell-types in others. We sought to analyze the clinical utility of a novel assay for putative leukemia stem cells in a large prospective cohort. The leukemic clone's most primitive hematopoietic cellular phenotype was prospectively identified in 109 newly-diagnosed acute myeloid leukemia patients, and analyzed against clinical risk groups and outcomes. Most (80/109) patients harbored CD34(+)CD38(-) leukemia cells. The CD34(+)CD38(-) leukemia cells in 47 of the 80 patients displayed intermediate aldehyde dehydrogenase expression, while normal CD34(+)CD38(-) hematopoietic stem cells expressed high levels of aldehyde dehydrogenase. In the other 33/80 patients, the CD34(+)CD38(-) leukemia cells exhibited high aldehyde dehydrogenase activity, and most (28/33, 85%) harbored poor-risk cytogenetics or FMS-like tyrosine kinase 3 internal tandem translocations. No CD34(+) leukemia cells could be detected in 28/109 patients, including 14/21 patients with nucleophosmin-1 mutations and 6/7 acute promyelocytic leukemia patients. The patients with CD34(+)CD38(-) leukemia cells with high aldehyde dehydrogenase activity manifested a significantly lower complete remission rate, as well as poorer event-free and overall survivals. The leukemic clone's most immature phenotype was heterogeneous with respect to CD34, CD38, and ALDH expression, but correlated with acute myeloid leukemia risk groups and outcomes. The strong clinical correlations suggest that the most immature phenotype detectable in the leukemia might serve as a biomarker for "clinically-relevant" leukemia stem cells. ClinicalTrials.gov: NCT01349972.

Regulation of human hematopoietic stem cell self-renewal by the microenvironment's control of retinoic acid signaling.

The high expression of aldehyde dehydrogenase 1, also known as retinaldehyde dehydrogenase, by hematopoietic stem cells (HSCs) suggests an important role for retinoic acid (RA) signaling in determining the fate of these cells. We found that primitive human bone marrow-derived CD34(+)CD38(-) cells not only highly express aldehyde dehydrogenase 1, but also the RA receptor α. Despite the up-regulation of early components of RA signaling, the downstream pathway remained inactive in the primitive CD34(+)CD38(-) cells. Primitive hematopoietic cells rapidly undergo terminal differentiation when cultured away from their microenvironment; however, we found that inhibition of RA signaling maintained their primitive phenotype and function, and promoted their self-renewal. HSCs reside in a complex microenvironment that enforces the balance between self-renewal and differentiation. The exact physiologic mechanisms by which the niche controls HSC fate remain elusive. The embryonic gonadal microenvironment has recently been shown to determine germ-cell fate by degrading RA through expression of the P450 retinoid-inactivating enzyme CYP26B1. We found that the bone marrow microenvironment similarly can control primitive hematopoietic cell fate via modulation of retinoid bioavailability. Accordingly, we found that bone marrow stromal cell CYP26 was also able to inactivate retinoids in serum, preventing RA signaling. Thus, primitive hematopoietic cells appear to be intrinsically programmed to undergo RA-mediated differentiation unless prevented from doing so by bone marrow niche CYP26. Modulation of RA signaling also holds promise for clinical HSC expansion, a prerequisite for the wide-scale use of these cells in regenerative medicine and gene therapy.

Preoperative predictors of pathological lymph node metastasis in patients with renal cell carcinoma undergoing retroperitoneal lymph node dissection.

PURPOSE: Patients with locally advanced renal cell carcinoma represent a subset that may benefit from retroperitoneal lymph node dissection. We identified preoperative clinical predictors of positive lymph nodes in patients with renal cell carcinoma without distant metastasis who underwent retroperitoneal lymph node dissection. MATERIALS AND METHODS: We retrospectively analyzed data on a consecutive cohort of 1,270 patients with cTany Nany M0 renal cell carcinoma who were treated at a single institution from 1993 to 2012. Multivariate analysis was performed to determine preoperative predictors of pathologically positive lymph nodes in patients who underwent retroperitoneal lymph node dissection. A nomogram was developed to predict the probability of lymph node metastasis. Overall, cancer specific and recurrence-free survival was estimated using the Kaplan-Meier Method. RESULTS: We identified 1,270 patients with renal cell carcinoma without distant metastasis who had (564) or did not have (706) retroperitoneal lymph node dissection performed. Of the 564 patients 131 (23%) and 433 (77%) had pN1 and pN0 disease, and 60 (37%) and 29 (7.2%) had cN1pN0 and cN0pN1 disease, respectively. ECOG PS, cN stage, local symptoms and lactate dehydrogenase were associated with nodal metastasis on multivariable analysis. A nomogram was developed with a C-index of 0.89 that demonstrated excellent calibration. Differences in overall, cancer specific and recurrence-free survival among pNx, pN0 and pN1 cases were statistically significant (p <0.001). CONCLUSIONS: Local symptoms, ECOG PS, cN stage and lactate dehydrogenase were independent predictors of lymph node metastasis in patients who underwent retroperitoneal lymph node dissection. Our predictive nomogram using these factors showed excellent discrimination and calibration.

A Sertoli cell-specific connexin43 knockout leads to altered interstitial connexin expression and increased Leydig cell numbers.

The Sertoli cell (SC)-specific knockout (KO) of connexin43 (Cx43) results in spermatogenic arrest at the level of spermatogonia and/or SC-only syndrome. Histology of the interstitial compartment suggests Leydig cell (LC) hyperplasia. Our aim has been to investigate possible effects of the SC-specific KO of Cx43 (SCCx43KO) on interstitial LC. We therefore counted LC via the optical dissector method (per microliter of testicular tissue and per testis) and found LC to be significantly increased in SCCx43KO(-/-) compared with wild-type mice. Semiquantitative western blot together with Cx43 and 3ß-hydroxysteroid dehydrogenase immunohistochemistry showed that Cx43 protein was significantly reduced and barely detectable in LC in adult SCCx43KO(-/-) mice. This reduction of Cx43 protein was accompanied by a reduction of Cx43 mRNA as analyzed by laser-assisted microdissection of interstitial cells and subsequent quantitative real-time polymerase chain reaction (PCR). Interestingly, Cx45, another recently detected connexin in LC, was also downregulated. Preliminary qualitative data of LC differentiation markers (Thb2, Hsd3b6) and a steroidogenic marker (Hsd17b3) obtained by reverse transcription plus PCR revealed no obvious differences. Thus, the loss of Cx43 in SC also provokes the downregulation of connexins in interstitial LC at the transcriptional and translational levels. Moreover, SCCx43KO leads to alterations in LC numbers. Despite these alterations, steroidogenesis seems not to be impaired. Further studies, including ultrastructural analysis of the tissue as well as quantitative examination of additional LC markers and testosterone, and functional in vitro experiments, should provide more information about LC differentiation and function in SCCx43KO(-/-) mice.

A clinically relevant population of leukemic CD34(+)CD38(-) cells in acute myeloid leukemia.

Relapse of acute myeloid leukemia (AML) is thought to reflect the failure of current therapies to adequately target leukemia stem cells (LSCs), the rare, resistant cells presumed responsible for maintenance of the leukemia and typically enriched in the CD34(+)CD38(-) cell population. Despite the considerable research on LSCs over the past 2 decades, the clinical significance of these cells remains uncertain. However, if clinically relevant, it is expected that LSCs would be enriched in minimal residual disease and predictive of relapse. CD34(+) subpopulations from AML patients were analyzed by flow cytometry throughout treatment. Sorted cell populations were analyzed by fluorescence in situ hybridization for leukemia-specific cytogenetic abnormalities (when present) and by transplantation into immunodeficient mice to determine self-renewal capacity. Intermediate (int) levels of aldehyde dehydrogenase (ALDH) activity reliably distinguished leukemic CD34(+)CD38(-) cells capable of engrafting immunodeficient mice from residual normal hematopoietic stem cells that exhibited relatively higher ALDH activity. Minimal residual disease detected during complete remission was enriched for the CD34(+)CD38(-)ALDH(int) leukemic cells, and the presence of these cells after therapy highly correlated with subsequent clinical relapse. ALDH activity appears to distinguish normal from leukemic CD34(+)CD38(-) cells and identifies those AML cells associated with relapse.