Metatranscriptomes generated by pyrosequencing hold significant potential for describing functional processes in complex microbial communities. Meeting this potential requires protocols that maximize ...mRNA recovery by reducing the relative abundance of ribosomal RNA, as well as systematic comparisons to identify methodological artifacts and test for reproducibility across data sets. Here, we implement a protocol for subtractive hybridization of bacterial rRNA (16S and 23S) that uses sample-specific probes and is applicable across diverse environmental samples. To test this method, rRNA-subtracted and unsubtracted transcriptomes were sequenced (454 FLX technology) from bacterioplankton communities at two depths in the oligotrophic open ocean, yielding 10 data sets representing approximately 350 Mbp. Subtractive hybridization reduced bacterial rRNA transcript abundance by 40-58%, increasing recovery of non-rRNA sequences up to fourfold (from 12% to 20% of total sequences to 40-49%). In testing this method, we established criteria for detecting sequences replicated artificially via pyrosequencing errors and identified such replicates as a significant component (6-39%) of total pyrosequencing reads. Following replicate removal, statistical comparisons of reference genes (identified via BLASTX to NCBI-nr) between technical replicates and between rRNA-subtracted and unsubtracted samples showed low levels of differential transcript abundance (<0.2% of reference genes). However, gene overlap between data sets was remarkably low, with no two data sets (including duplicate runs from the same pyrosequencing library template) sharing greater than 17% of unique reference genes. These results indicate that pyrosequencing captures a small subset of total mRNA diversity and underscores the importance of reliable rRNA subtraction procedures to enhance sequencing coverage across the functional transcript pool.
Background
Anastomotic leak after esophagectomy is associated with significant morbidity and mortality. Our institution began performing laparoscopic gastric ischemic preconditioning (LGIP) with ...ligation of the left gastric and short gastric vessels prior to esophagectomy in all patients presenting with resectable esophageal cancer. We hypothesized that LGIP may decrease the incidence and severity of anastomotic leak.
Methods
Patients were prospectively evaluated following the universal application of LGIP prior to esophagectomy protocol in January 2021 until August 2022. Outcomes were compared with patients who underwent esophagectomy without LGIP from a prospectively maintained database from 2010 to 2020.
Results
We compared 42 patients who underwent LGIP followed by esophagectomy with 222 who underwent esophagectomy without LGIP. Age, sex, comorbidities, and clinical stage were similar between groups. Outpatient LGIP was generally well tolerated, with one patient experiencing prolonged gastroparesis. Median time from LGIP to esophagectomy was 31 days. Mean operative time and blood loss were not significantly different between groups. Patients who underwent LGIP were significantly less likely to develop an anastomotic leak following esophagectomy (7.1% vs. 20.7%,
p
= 0.038). This finding persisted on multivariate analysis odds ratio (OR) 0.17, 95% confidence interval (CI) 0.03–0.42,
p
= 0.029. The occurrence of any post-esophagectomy complication was similar between groups (40.5% vs. 46.0%,
p
= 0.514), but patients who underwent LGIP had shorter length of stay 10 (9–11) vs. 12 (9–15),
p
= 0.020.
Conclusions
LGIP prior to esophagectomy is associated with a decreased risk of anastomotic leak and length of hospital stay. Further, multi-institutional studies are warranted to confirm these findings.
Increasing evidence suggests that hysterectomy to treat uterine fibroids (UFs), even with ovarian conservation (OC), is associated with a 33% increased risk of coronary artery disease (CAD). We ...sought to compare the cost-effectiveness of various treatment approaches for UFs to understand the trade-offs among development of CAD vs new fibroids.
We developed a Markov model to include women with UFs who no longer desired pregnancy. The outcomes of interest were quality-adjusted life-years (QALYs) and total treatment costs. We conducted sensitivity analyses to test the effect of uncertain model inputs.
Health system perspective.
A hypothetical cohort of 10 000 40-year-old women.
Myomectomy, hysterectomy with OC, and hysterectomy without OC.
Myomectomy was the best-value strategy, costing US$528 217 and providing 19.38 QALYs. Neither hysterectomy with OC nor hysterectomy without OC was found to be cost-effective, assuming a willingness-to-pay threshold of $100 000 per QALY gain as hysterectomy with OC provided more benefit than myomectomy at an average cost of $613 144 to gain one additional QALY. The sensitivity analyses showed that if the risk of new symptomatic UFs that required treatment after myomectomy was more than 13%, annually (base case, 3.6%), or the quality of life after myomectomy was less than 0.815 (base case, 0.834), then myomectomy would no longer be cost-effective, under a willingness-to-pay amount of US$100 000.
Myomectomy is an optimal treatment of UFs compared with hysterectomy among women aged 40 years. The increased risk of CAD after hysterectomy and its associated costs and the effects on morbidity and quality of life made hysterectomy a costlier and less effective long-term strategy.
Knowledge of the neural mechanisms underlying increased disease burden in anxiety disorders that is unaccounted for by individual categorical diagnoses could lead to improved clinical care. Here, we ...tested the utility of a joint functional magnetic resonance imaging–electroencephalography neurobiological profile characterized by overvaluation of negative stimuli (amygdala) in combination with blunted elaborated processing of these same stimuli (the late positive potential LPP, an event-related potential) in predicting increased psychopathology across a 2-year period in people with anxiety disorders.
One hundred ten participants (64 female, 45 male, 1 other) including 78 participants with phobias who varied in the extent of their internalizing comorbidity and 32 participants who were free from psychopathology viewed negative and neutral pictures during separate functional magnetic resonance imaging blood oxygen level–dependent and electroencephalogram recordings. Dysphoria was assessed at baseline and 2 years later.
Participants with both heightened amygdala activation and blunted LPPs to negative pictures showed the greatest increases in dysphoria 2 years later. Cross-sectionally, participants with higher comorbidity load (≥2 additional diagnoses, n = 34) showed increased amygdala activation to negative pictures compared with participants with lower comorbidity load (≤1 additional diagnosis, n = 44) and compared with participants free from psychopathology. In addition, high comorbid participants showed reduced LPPs to negative pictures compared with low comorbid participants.
Heightened amygdala in response to negative stimuli in combination with blunted LPPs could indicate overvaluation of threatening stimuli in the absence of elaborated processing that might otherwise help regulate threat responding. This brain profile could underlie the worsening and maintenance of internalizing psychopathology over time.
There is growing consensus that intervention and treatment of Huntington disease (HD) should occur at the earliest stage possible. Various early-intervention methods for this fatal neurodegenerative ...disease have been identified, but preventive clinical trials for HD are limited by a lack of knowledge of the natural history of the disease and a dearth of appropriate outcome measures. Objectives of the current study are to document the natural history of premanifest HD progression in the largest cohort ever studied and to develop a battery of imaging and clinical markers of premanifest HD progression that can be used as outcome measures in preventive clinical trials. Neurobiological predictors of Huntington's disease is a 32-site, international, observational study of premanifest HD, with annual examination of 1013 participants with premanifest HD and 301 gene-expansion negative controls between 2001 and 2012. Findings document 39 variables representing imaging, motor, cognitive, functional, and psychiatric domains, showing different rates of decline between premanifest HD and controls. Required sample size and models of premanifest HD are presented to inform future design of clinical and preclinical research. Preventive clinical trials in premanifest HD with participants who have a medium or high probability of motor onset are calculated to be as resource-effective as those conducted in diagnosed HD and could interrupt disease 7-12 years earlier. Methods and measures for preventive clinical trials in premanifest HD more than a dozen years from motor onset are also feasible. These findings represent the most thorough documentation of a clinical battery for experimental therapeutics in stages of premanifest HD, the time period for which effective intervention may provide the most positive possible outcome for patients and their families affected by this devastating disease.
Uterine leiomyomas are common hormone-responsive neoplasms that frequently cause heavy menstrual bleeding, anemia, pelvic pressure, pain, and adverse reproductive outcomes. In this overview, the ...efficacy and safety of oral gonadotropin-releasing hormone (GnRH) antagonists, co-administered with menopausal replacement-level steroid hormones or used at doses to avoid complete hypothalamic suppression, are reviewed for the management of uterine leiomyomas. Oral GnRH antagonists provide rapid suppression of sex steroids and avoid the initial steroidal flare and resultant temporary worsening of symptoms typically seen with parenteral GnRH agonists. Oral GnRH antagonists are effective in reducing leiomyoma-associated heavy menstrual bleeding, with high rates of amenorrhea and improved anemia and leiomyoma-associated pain, and providing modest reduction in uterine volume when used in combination with menopausal replacement-level steroid hormones. This add-back therapy can reduce hypogonadal side effects, including hot flushes and bone mineral density loss, close to levels seen with placebo therapy. Currently, both elagolix 300 mg twice daily with once-daily estradiol (1 mg) and norethindrone (0.5 mg) and relugolix 40 mg once daily with estradiol (1 mg) and norethindrone (0.5 mg) combination therapy are approved for leiomyoma treatment by the U.S. Food and Drug Administration. Linzagolix is under investigation in the United States but approved at two does with and without steroid hormones in the European Union. The efficacy of these agents appears to be robust over a wide spectrum of clinical presentations, demonstrating that worse disease parameters at baseline do not appear to inhibit efficacy. Across clinical trials, participants largely reflected the population of individuals affected by uterine leiomyomas.
Uterine fibroids are one of the most common neoplasms found among women globally, with a prevalence of approximately 11 million women in the United States alone. The morbidity of this common disease ...is significant because it is the leading cause of hysterectomy and causes significant functional impairment for women of reproductive age. Factors including age, body mass index, race, ethnicity, menstrual blood loss, fibroid location, and uterine and fibroid volume influence the incidence of fibroids and severity of symptoms. Elagolix is an oral gonadotropin-releasing hormone receptor antagonist that competitively inhibits pituitary gonadotropin-releasing hormone receptor activity and suppresses the release of gonadotropins from the pituitary gland, resulting in dose-dependent suppression of ovarian sex hormones, follicular growth, and ovulation. In Elaris Uterine Fibroids 1 and Uterine Fibroids 2, 2 replicate multicenter, double-blind, randomized, placebo-controlled, phase 3 studies, treatment of premenopausal women with elagolix with hormonal add-back therapy demonstrated reduction in heavy menstrual bleeding associated with uterine fibroids.
This analysis aimed to evaluate the safety and efficacy of elagolix (300 mg twice a day) with add-back therapy (1 mg estradiol/0.5 mg norethindrone acetate once a day) in reducing heavy menstrual bleeding associated with uterine fibroids in various subgroups of women over 6 months of treatment.
Data were pooled from Elaris Uterine Fibroid-1 and Uterine Fibroid-2 studies, which evaluated premenopausal women (18–51 years) with heavy menstrual bleeding (>80 mL menstrual blood loss per cycle, alkaline hematin methodology) and ultrasound-confirmed uterine fibroid diagnosis. Subgroups analyzed included age, body mass index, race, ethnicity, baseline menstrual blood loss, fibroid location, and uterine and primary fibroid volume (largest fibroid identified by ultrasound). The primary endpoint was the proportion of women with <80 mL menstrual blood loss during the final month and ≥50% menstrual blood loss reduction from baseline to final month. Secondary and other efficacy endpoints included mean change in menstrual blood loss from baseline to final month, amenorrhea, symptom severity, and health-related quality of life. Adverse events and other safety endpoints were monitored.
The overall pooled Elaris Uterine Fibroid-1 and Uterine Fibroid-2 population was typical of women with fibroids, with a mean age of 42.4 (standard deviation, 5.4) years and a mean body mass index of 33.6 (standard deviation, 7.3) kg/m2 and 67.6% of participants being black or African American women. A wide range of baseline uterine and fibroid volumes and menstrual blood loss were also represented in the overall pooled study population. In all subgroups, the proportion of responders to the primary endpoint, mean change in menstrual blood loss, amenorrhea, reduction in symptom severity, and improvement in health-related quality of life were clinically meaningfully greater for women who received elagolix with add-back therapy than those who received placebo and consistent with the overall pooled study population for the primary endpoint (72.2% vs 9.3%), mean change in menstrual blood loss (−172.5 mL vs −0.8 mL), amenorrhea (50.4% vs 4.5%), symptom severity (−37.1 vs −9.2), and health-related quality of life score (39.9 vs 8.9). Adverse events by subgroup were consistent with the overall pooled study population.
Elagolix with hormonal add-back therapy was effective in reducing heavy menstrual bleeding associated with uterine fibroids independent of age, body mass index, race, ethnicity, baseline menstrual blood loss, fibroid location, and uterine and primary fibroid volume.
To assess the effect of once-daily relugolix combination therapy (relugolix-CT: relugolix 40 mg, estradiol 1 mg, and norethindrone acetate 0.5 mg) compared with placebo on moderate-to-severe pain in ...women with uterine leiomyomas and heavy menstrual bleeding.
Two replicate, multinational, double-blind, 24-week, randomized, phase 3 studies (LIBERTY 1 and 2) were conducted in premenopausal women with uterine leiomyoma-associated heavy menstrual bleeding (80 mL or greater per cycle for two cycles or 160 mL or greater during one cycle). A predefined secondary objective was to determine the effect of relugolix-CT on moderate-to-severe uterine leiomyoma-associated pain in the pain subpopulation (women with maximum pain scores of 4 or higher on the 0-10 numerical rating scale at baseline, with pain score reporting compliance of 80% (ie, 28 days or more over the last 35 days of treatment). This key secondary endpoint was defined as the proportion of women achieving minimal-to-no uterine leiomyoma-associated pain (maximum numerical rating scale score 1 or lower) at week 24; menstrual and nonmenstrual pain were evaluated in prespecified secondary analyses. Treatment comparisons were performed in the pooled LIBERTY 1 and 2 pain subpopulation using the Cochran-Mantel-Haenszel test stratified by baseline menstrual blood loss volume.
Across both trials, 509 women were randomized to relugolix-CT or placebo (April 2017-December 2018). Of these, 277 (54.4%) met pain subpopulation requirements. With relugolix-CT, 45.2% (95% CI 36.4-54.3) of women achieved minimal-to-no pain compared with 13.9% (95% CI 8.8-20.5) with placebo (nominal P<.001). The proportions of women with minimal-to-no pain during menstrual days and during nonmenstrual days were significantly higher with relugolix-CT (65.0% 95% CI 55.6-73.5 and 44.6% 95% CI 32.3-57.5, respectively) compared with placebo (19.3% 95% CI 13.2-26.7, nominal P<.001, and 21.6% 95% CI 12.9-32.7, nominal P=.004, respectively).
Over 24 weeks, relugolix-CT significantly reduced moderate-to-severe uterine leiomyoma-associated pain with a more pronounced effect on menstrual pain. These data support that relugolix-CT had clinically meaningful effects on women's experience of uterine leiomyoma-associated pain.
ClinicalTrials.gov: LIBERTY 1, NCT03049735; LIBERTY 2, NCT03103087.
Myovant Sciences GmbH.
To assess several measures of the long-term outcome of magnetic resonance-guided focused ultrasound surgery for symptomatic uterine leiomyomata.
Data on 359 women completing 24-month follow-up in all ...clinical trials of magnetic resonance-guided focused ultrasound surgery for uterine leiomyomata were analyzed. Quality of life outcomes, measured by the symptom severity score of the Uterine Fibroid Symptoms Quality Of Life Questionnaire were assessed for 24 months after treatment. Clinical endpoints, including uterine shrinkage, the need for additional leiomyoma treatment, and the time to additional leiomyoma treatment, were all assessed. The nonperfused volume ratio after treatment, calculated from the gadolinium-enhanced magnetic resonance imaging after treatment and the best measure of tissue necrosis after treatment, was used to assess outcome based on completeness of leiomyoma ablation.
Women undergoing magnetic resonance-guided focused ultrasound surgery for symptomatic uterine leiomyomata have durable symptom relief, as measured by the symptom severity score at 24 months, with significantly greater improvement with more complete ablation (P<.001). Survival analysis demonstrates a significant reduction in the percentage of women undergoing additional leiomyoma treatment (P=.001) in women in the high nonperfused volume group. The mean shrinkage and mean residual nonperfused volume ratio are both significantly above zero at 6 months in the high nonperfused volume group (P<.001). The incidence of adverse events is low. However, for women with minimal treatment, the risk of additional procedures is high.
Magnetic resonance-guided focused ultrasound surgery is an effective treatment for uterine leiomyomata and results in sustained symptomatic relief.
III.
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