Complement in the Brain: Contributions to Neuroprotection, Neuronal Plasticity, and Neuroinflammation.

The complement system is an ancient collection of proteolytic cascades with well-described roles in regulation of innate and adaptive immunity. With the convergence of a revolution in complement-directed clinical therapeutics, the discovery of specific complement-associated targetable pathways in the central nervous system, and the development of integrated multi-omic technologies that have all emerged over the last 15 years, precision therapeutic targeting in Alzheimer disease and other neurodegenerative diseases and processes appears to be within reach. As a sensor of tissue distress, the complement system protects the brain from microbial challenge as well as the accumulation of dead and/or damaged molecules and cells. Additional more recently discovered diverse functions of complement make it of paramount importance to design complement-directed neurotherapeutics such that the beneficial roles in neurodevelopment, adult neural plasticity, and neuroprotective functions of the complement system are retained.

Molecular contribution to embryonic aneuploidy and karyotypic complexity in initial cleavage divisions of mammalian development.

Embryonic aneuploidy is highly complex, often leading to developmental arrest, implantation failure or spontaneous miscarriage in both natural and assisted reproduction. Despite our knowledge of mitotic mis-segregation in somatic cells, the molecular pathways regulating chromosome fidelity during the error-prone cleavage-stage of mammalian embryogenesis remain largely undefined. Using bovine embryos and live-cell fluorescent imaging, we observed frequent micro-/multi-nucleation of mis-segregated chromosomes in initial mitotic divisions that underwent unilateral inheritance, re-fused with the primary nucleus or formed a chromatin bridge with neighboring cells. A correlation between a lack of syngamy, multipolar divisions and asymmetric genome partitioning was also revealed, and single-cell DNA-seq showed propagation of primarily non-reciprocal mitotic errors. Depletion of the mitotic checkpoint protein BUB1B (also known as BUBR1) resulted in similarly abnormal nuclear structures and cell divisions, as well as chaotic aneuploidy and dysregulation of the kinase-substrate network that mediates mitotic progression, all before zygotic genome activation. This demonstrates that embryonic micronuclei sustain multiple fates, provides an explanation for blastomeres with uniparental origins, and substantiates defective checkpoints and likely other maternally derived factors as major contributors to the karyotypic complexity afflicting mammalian preimplantation development.

A systematic review and meta-analysis on the transcriptomic signatures in alcohol use disorder.

Currently available clinical treatments on alcohol use disorder (AUD) exhibit limited efficacy and new druggable targets are required. One promising approach to discover new molecular treatment targets involves the transcriptomic profiling of brain regions within the addiction neurocircuitry, utilizing animal models and postmortem brain tissue from deceased patients with AUD. Unfortunately, such studies suffer from large heterogeneity and small sample sizes. To address these limitations, we conducted a cross-species meta-analysis on transcriptome-wide data obtained from brain tissue of patients with AUD and animal models. We integrated 36 cross-species transcriptome-wide RNA-expression datasets with an alcohol-dependent phenotype vs. controls, following the PRISMA guidelines. In total, we meta-analyzed 964 samples - 502 samples from the prefrontal cortex (PFC), 282 nucleus accumbens (NAc) samples, and 180 from amygdala (AMY). The PFC had the highest number of differentially expressed genes (DEGs) across rodents, monkeys, and humans. Commonly dysregulated DEGs suggest conserved cross-species mechanisms for chronic alcohol consumption/AUD comprising MAPKs as well as STAT, IRF7, and TNF. Furthermore, we identified numerous unique gene sets that might contribute individually to these conserved mechanisms and also suggest novel molecular aspects of AUD. Validation of the transcriptomic alterations on the protein level revealed interesting targets for further investigation. Finally, we identified a combination of DEGs that are commonly regulated across different brain tissues as potential biomarkers for AUD. In summary, we provide a compendium of genes that are assessable via a shiny app, and describe signaling pathways, and physiological and cellular processes that are altered in AUD that require future studies for functional validation.

Recurrence and Survival After Minimally Invasive and Open Esophagectomy for Esophageal Cancer: A Post Hoc Analysis of the Ensure Study.

OBJECTIVE: To determine the impact of operative approach [open (OE), hybrid minimally invasive (HMIE), and total minimally invasive (TMIE) esophagectomy] on operative and oncologic outcomes for patients treated with curative intent for esophageal and junctional cancer. BACKGROUND: The optimum oncologic surgical approach to esophageal and junctional cancer is unclear. METHODS: This secondary analysis of the European multicenter ENSURE study includes patients undergoing curative-intent esophagectomy for cancer between 2009 and 2015 across 20 high-volume centers. Primary endpoints were disease-free survival (DFS) and the incidence and location of disease recurrence. Secondary endpoints included among others R0 resection rate, lymph node yield, and overall survival (OS). RESULTS: In total, 3199 patients were included. Of these, 55% underwent OE, 17% HMIE, and 29% TMIE. DFS was independently increased post-TMIE [hazard ratio (HR): 0.86 (95% CI: 0.76-0.98), P = 0.022] compared with OE. Multivariable regression demonstrated no difference in absolute locoregional recurrence risk according to the operative approach [HMIE vs OE, odds ratio (OR): 0.79, P = 0.257; TMIE vs OE, OR: 0.84, P = 0.243]. The probability of systemic recurrence was independently increased post-HMIE (OR: 2.07, P = 0.031), but not TMIE (OR: 0.86, P = 0.508). R0 resection rates ( P = 0.005) and nodal yield ( P < 0.001) were independently increased after TMIE, but not HMIE ( P = 0.424; P = 0.512) compared with OE. OS was independently improved following both HMIE (HR: 0.79, P = 0.009) and TMIE (HR: 0.82, P = 0.003) as compared with OE. CONCLUSION: In this European multicenter study, TMIE was associated with improved surgical quality and DFS, whereas both TMIE and HMIE were associated with improved OS as compared with OE for esophageal cancer.

Evolution in Laparoscopic Gastrectomy From a Randomized Controlled Trial Through National Clinical Practice.

OBJECTIVE: To examine the influence of the LOGICA RCT (randomized controlled trial) upon the practice and outcomes of laparoscopic gastrectomy within the Netherlands. BACKGROUND: Following RCTs the dissemination of complex interventions has been poorly studied. The LOGICA RCT included 10 Dutch centers and compared laparoscopic to open gastrectomy. METHODS: Data were obtained from the Dutch Upper Gastrointestinal Cancer Audit (DUCA) on all gastrectomies performed in the Netherlands (2012-2021), and the LOGICA RCT from 2015 to 2018. Multilevel multivariable logistic regression analyses were performed to assess the effect of laparoscopic versus open gastrectomy upon clinical outcomes before, during, and after the LOGICA RCT. RESULTS: Two hundred eleven patients from the LOGICA RCT (105 open vs 106 laparoscopic) and 4131 patients from the DUCA data set (1884 open vs 2247 laparoscopic) were included. In 2012, laparoscopic gastrectomy was performed in 6% of patients, increasing to 82% in 2021. No significant effect of laparoscopic gastrectomy on postoperative clinical outcomes was observed within the LOGICA RCT. Nationally within DUCA, a shift toward a beneficial effect of laparoscopic gastrectomy upon complications was observed, reaching a significant reduction in overall [adjusted odds ratio (aOR):0.62; 95% CI: 0.46-0.82], severe (aOR: 0.64; 95% CI: 0.46-0.90) and cardiac complications (aOR: 0.51; 95% CI: 0.30-0.89) after the LOGICA trial. CONCLUSIONS: The wider benefits of the LOGICA trial included the safe dissemination of laparoscopic gastrectomy across the Netherlands. The robust surgical quality assurance program in the design of the LOGICA RCT was crucial to facilitate the national dissemination of the technique following the trial and reducing potential patient harm during surgeons learning curve.

Optimal Treatment Strategies for cT2 Staged Adenocarcinoma of the Esophagus and the Gastroesophageal Junction: A Multinational, High-volume Center Retrospective Cohort Analysis.

OBJECTIVE: A multi-national high-volume center study was undertaken to evaluate outcomes after primary surgery (PS) or neoadjuvant treatment followed by surgery (NAT/S) in cT2 staged adenocarcinomas of the esophagus (EAC) and gastroesophageal junction (GEJ). BACKGROUND: Optimal treatment approach with either NAT/S or PS for clinically staged cT2cNany or cT2N0 EAC and GEJ remains unknown due to the lack of randomized controlled trials. METHODS: Retrospective analysis of prospectively maintained databases from ten centers was performed. Between 01/2012-08/2023 645 patients who fulfilled inclusion criteria of GEJ Siewert type I, II or EAC with cT2 status at diagnosis underwent PS or NAT/S with curative intent. Primary endpoint was overall survival (OS). RESULTS: In the cT2cNany cohort 192 patients (29.8%) underwent PS and 453 (70.2%) underwent NAT/S. In all cT2cN0 patients (n=333), NAT/s remained the more frequent treatment (56.2%). Patients undergoing PS were in both cT2 cohorts older (P<0.001) and had a higher ASA classification (P<0.05). R0 resection showed no differences between NAT/S and PS in both cT2 cohorts (P>0.4).Median OS was 51.0 months in the PS group (95% CI 31.6-70.4) versus 114.0 months (95% CI 53.9-174.1) in the NAT/S group (P=0.003) of cT2cNany patients. For cT2cN0 patients NAT/S was associated with longer OS (P=0.002) and disease-free survival (DFS) (P=0.001). After propensity score matching of cT2N0 patients, survival benefit for NAT/S remained (P=0.004). Histopathology showed that 38.1% of cT2cNany and 34.2% of cT2cN0 patients were understaged. CONCLUSIONS: Due to unreliable identification of cT2N0 disease, all patients should be offered a multimodal therapeutic approach.

The Prognostic Impact of Minimally Invasive Esophagectomy on Survival After Esophagectomy Following a Delayed Interval After Chemoradiotherapy: A Secondary Analysis of the DICE Study.

OBJECTIVE: To evaluate prognostic differences between minimally invasive esophagectomy (MIE) and open esophagectomy (OE) in patients with surgery after a prolonged interval (>12 wk) following chemoradiotherapy (CRT). BACKGROUND: Previously, we established that a prolonged interval after CRT before esophagectomy was associated with poorer long-term survival. METHODS: This was an international multicenter cohort study involving 17 tertiary centers, including patients who received CRT followed by surgery between 2010 and 2020. Patients undergoing MIE were defined as thoracoscopic and laparoscopic approaches. RESULTS: A total of 428 patients (145 MIE and 283 OE) had surgery between 12 weeks and 2 years after CRT. Significant differences were observed in American Society of Anesthesiologists grade, radiation dose, clinical T stage, and histologic subtype. There were no significant differences between the groups in age, sex, body mass index, pathologic T or N stage, resection margin status, tumor location, surgical technique, or 90-day mortality. Survival analysis showed MIE was associated with improved survival in univariate ( P =0.014), multivariate analysis after adjustment for smoking, T and N stage, and histology (HR=1.69; 95% CI: 1.14-2.5) and propensity-matched analysis ( P =0.02). Further subgroup analyses by radiation dose and interval after CRT showed survival advantage for MIE in 40 to 50 Gy dose groups (HR=1.9; 95% CI: 1.2-3.0) and in patients having surgery within 6 months of CRT (HR=1.6; 95% CI: 1.1-2.2). CONCLUSIONS: MIE was associated with improved overall survival compared with OE in patients with a prolonged interval from CRT to surgery. The mechanism for this observed improvement in survival remains unknown, with potential hypotheses including a reduction in complications and improved functional recovery after MIE.

Paravertebral versus EPidural Analgesia in Minimally Invasive Esophageal ResectioN (PEPMEN): A Randomized Controlled Multicenter Trial.

OBJECTIVE: To compare quality of recovery in patients receiving epidural or paravertebral analgesia for minimally invasive esophagectomy (MIE). SUMMARY BACKGROUND DATA: Paravertebral analgesia may be a promising alternative to epidural analgesia, avoiding potential side effects and improving postoperative recovery. METHODS: This randomized controlled superiority trial was conducted across four Dutch centers with esophageal cancer patients scheduled for transthoracic MIE with intrathoracic anastomosis, randomizing patients to receive either epidural or paravertebral analgesia. The primary outcome was Quality of Recovery (QoR-40) on the third postoperative day (POD). Secondary outcomes included quality of life, postoperative pain, opioid consumption, inotropic/vasopressor medication use, hospital stay, complications, readmission, and mortality. RESULTS: From December 2019 to February 2023, 192 patients were included: 94 received epidural and 98 paravertebral analgesia. QoR-40 score on POD3 was not different between groups (mean difference 3.7, 95%CI -2.3 to 9.7; P=0.268). Epidural patients had significant higher QoR-40 scores on POD1 and 2 (mean difference 7.7, 95%CI 2.3-13.1; P=0.018 and mean difference 7.3, 95%CI 1.9-12.7; P=0.020) and lower pain scores (median 1 versus 2; P=<0.001 and median 1 versus 2; P=0.033). More epidural patients required vasopressor medication on POD1 (38.3% versus 13.3%; P<0.001). Urinary catheters were removed earlier in the paravertebral group (median POD3 versus 4; P=<0.001). No significant differences were found in postoperative complications or hospital/Intensive Care Unit stay. CONCLUSIONS: This randomized controlled trial did not demonstrate superiority of paravertebral over epidural analgesia regarding quality of recovery on POD3 after MIE. Both techniques are effective and can be offered in clinical practice.

Transcriptional profiling of mucus production in rhesus macaque endocervical cells under hormonal regulation.

OBJECTIVE: Endocervical mucus production is a key regulator of fertility throughout the menstrual cycle. With cycle-dependent variability in mucus quality and quantity, cervical mucus can either facilitate or block sperm ascension into the upper female reproductive tract. This study seeks to identify genes involved in the hormonal regulation of mucus production, modification, and regulation through profiling the transcriptome of endocervical cells from the non-human primate, the rhesus macaque (Macaca mulatta). INTERVENTION: We treated differentiated primary endocervical cultures with estradiol (E2) and progesterone (P4) to mimic peri-ovulatory and luteal-phase hormonal changes. Using RNA-sequencing, we identified differential expression of gene pathways and mucus producing and modifying genes in cells treated with E2 compared to hormone-free conditions and E2 compared to E2-primed cells treated with P4. MAIN OUTCOME MEASURES: We pursued differential gene expression analysis on RNA-sequenced cells. Sequence validation was done using qPCR. RESULTS: Our study identified 158 genes that show significant differential expression in E2-only conditions compared to hormone-free control, and 250 genes that show significant differential expression in P4-treated conditions compared to E2-only conditions. From this list, we found hormone-induced changes in transcriptional profiles for genes across several classes of mucus production, including ion channels and enzymes involved in post-translational mucin modification that have not previously been described as hormonally regulated. CONCLUSION: Our study is the first to use an in vitro culture system to create an epithelial-cell specific transcriptome of the endocervix. As a result, our study identifies new genes and pathways altered by sex-steroids in cervical mucus production.

Efficacy of SPR720 in murine models of non-tuberculous mycobacterial pulmonary infection.

BACKGROUND: Non-tuberculous mycobacterial pulmonary disease (NTM-PD) is increasing worldwide, with Mycobacterium avium complex (MAC) and Mycobacterium abscessus as the predominant pathogens. Current treatments are poorly tolerated and modestly effective, highlighting the need for new treatments. SPR719, the active moiety of the benzimidazole prodrug SPR720, inhibits the ATPase subunits of DNA gyrase B, a target not exploited by current antibiotics, and therefore, no cross-resistance is expected with standard-of-care (SOC) agents. OBJECTIVES: To evaluate the in vitro activity of SPR719 against MAC and M. abscessus clinical isolates, including those resistant to SOC agents, and in vivo efficacy of SPR720 in murine non-tuberculous mycobacteria (NTM) pulmonary infection models. METHODS: NTM isolates were tested for susceptibility to SPR719. Chronic C3HeB/FeJ and severe combined immunodeficient murine models of pulmonary infection were used to assess efficacy of SPR720 against MAC and M. abscessus, respectively. RESULTS: SPR719 was active against MAC (MIC90, 2 mg/L) and M. abscessus (MIC90, 4 mg/L) clinical isolates. Efficacy of SPR720 was demonstrated against MAC pulmonary infection, both as a monotherapy and in combination with SOC agents. SPR720 monotherapy exhibited dose-dependent reduction in bacterial burden, with the largest reduction observed when combined with clarithromycin and ethambutol. Efficacy of SPR720 was also demonstrated against M. abscessus pulmonary infection where monotherapy exhibited a dose-dependent reduction in bacterial burden with further reductions detected when combined with SOC agents. CONCLUSIONS: In vitro activity of SPR720 against common NTM pathogens and efficacy in murine infections warrant the continued clinical evaluation of SPR720 as a new oral option for the treatment of NTM-PD.