DNA mismatch and damage patterns revealed by single-molecule sequencing.

Mutations accumulate in the genome of every cell of the body throughout life, causing cancer and other diseases1,2. Most mutations begin as nucleotide mismatches or damage in one of the two strands of the DNA before becoming double-strand mutations if unrepaired or misrepaired3,4. However, current DNA-sequencing technologies cannot accurately resolve these initial single-strand events. Here we develop a single-molecule, long-read sequencing method (Hairpin Duplex Enhanced Fidelity sequencing (HiDEF-seq)) that achieves single-molecule fidelity for base substitutions when present in either one or both DNA strands. HiDEF-seq also detects cytosine deamination-a common type of DNA damage-with single-molecule fidelity. We profiled 134 samples from diverse tissues, including from individuals with cancer predisposition syndromes, and derive from them single-strand mismatch and damage signatures. We find correspondences between these single-strand signatures and known double-strand mutational signatures, which resolves the identity of the initiating lesions. Tumours deficient in both mismatch repair and replicative polymerase proofreading show distinct single-strand mismatch patterns compared to samples that are deficient in only polymerase proofreading. We also define a single-strand damage signature for APOBEC3A. In the mitochondrial genome, our findings support a mutagenic mechanism occurring primarily during replication. As double-strand DNA mutations are only the end point of the mutation process, our approach to detect the initiating single-strand events at single-molecule resolution will enable studies of how mutations arise in a variety of contexts, especially in cancer and ageing.

Personal genital satisfaction is associated with differences in perception of male and female sexual anatomy and function.

BACKGROUND: Prior studies primarily of men correlated low personal genital satisfaction (PGS) with decreased sexual activity; however, the association between PGS and genital anatomy perceptions is unknown, and there is a paucity of studies examining women. AIM: We assessed the relationship between genital satisfaction, survey respondent sexual activity, and perceptions of anatomy and function. METHODS: A 54-item REDCap survey was distributed to any-gendered volunteers ≥18 years of age through ResearchMatch from January to March 2023. Responses were split into (1) high PGS and (2) low PGS. Analysis was performed using chi-square tests on survey responses and a Mann Whitney U test on median satisfaction level. OUTCOMES: Outcomes were genital anatomy perceptions, sexual activity, and respondents' PGS. RESULTS: Of the 649 respondents who started the survey, 560 (86.3%) completed it. Median PGS was 7 of 10, forming subgroups of high (≥7 of 10) satisfaction (n = 317 of 560 [56.6%]) and low (<7 of 10) satisfaction (n = 243 of 560 [43.4%]). The mean age was 45.8 ± 16.8 years, and demographics were notable for 72.1% women (n = 404 of 560), 83.2% White (n = 466 of 560), 47.9% married (n = 268 of 560), and 75.5% bachelor's degree holders (n = 423 of 560). Comparing high- and low-PGS groups, more low-PGS respondents felt normal flaccid penis length to be <2 inches (11.1% vs 5.1%; P = .008). High-PGS respondents more often responded that it is normal for women to have orgasms over half the time (20.8% vs 13.2%; P = .0002) or to identify as being sexually active (81.1% vs 71.6%; P = .008). Women were more likely than men to report larger normal testicle sizes as 60.1 to 90 mL (24.5% vs 10.3%; P < .0001), whereas more men felt that normal testicle size was 7 to 15 mL (26.3% vs 11.4%; P < .0001). Orgasm length perceptions also differed: more women felt female orgasm length was 2.6 to 5 seconds (36.6% vs 16.7%; P < .0001), and more men believed female orgasms to be longer, at 7.6 to 10 seconds (29.5% vs 17.3%; P = .002), 10.1 to 12.5 seconds (11.5% vs 5.2%; P = .0008), and >12.5 seconds (12.2% vs 5.7%; P = .009). Respondents' views on their genitalia differed by gender, with women more likely to feel that their genitals are normal compared with men (89.4% vs 75.0%; P < .0001). CLINICAL IMPLICATIONS: PGS may be a useful screening tool given its association with sexual activity. STRENGTHS AND LIMITATIONS: Our large-scale survey assesses public perceptions of genital anatomy and function. Limitations include a lack of gender nonbinary perceptions. CONCLUSION: Gender and PGS interact with perceptions of male anatomy and female sexual activity, and the frequency of sexual activity was higher among high-PGS respondents; however, the direction of these interactions remains unclear and requires future causal analysis.

Testosterone therapy in females is not associated with increased cardiovascular or breast cancer risk: a claims database analysis.

BACKGROUND: Testosterone therapy (TTh) has been shown to improve libido in women with sexual dysfunction, but its utilization has been limited due to concern for cardiovascular events and past studies reporting highly variable results. AIM: To assess the association of TTh in women with major adverse cardiac events (MACEs), including heart attack, stroke, or death, using a large database. METHODS: The TriNetX Diamond Network was queried from 2009 to 2022. Our study cohort included adult females with ≥3 systemic testosterone prescriptions within a year. Our control cohort excluded females with any testosterone prescriptions, polycystic ovary syndrome, or androgen excess. Both cohorts excluded females with prior heart failure, unstable angina, intersex surgery (female to male), personal history of sex reassignment, or gender identity disorders. Propensity matching between the cohorts was performed. A subanalysis by age was conducted (18-55 and >55 years). OUTCOMES: We evaluated the association of TTh to the following: MACE, upper or lower emboli or deep vein thrombosis (DVT), pulmonary embolism (PE), breast neoplasm, and hirsutism within 3 years of TTh. RESULTS: When compared with propensity-matched controls, adult females with TTh had a lower risk of MACE (risk ratio [RR], 0.64; 95% CI, 0.51-0.81), DVT (RR, 0.61; 95% CI, 0.42-0.90), PE (RR, 0.48; 95% CI, 0.28-0.82), and malignant breast neoplasm (RR, 0.48; 95% CI, 0.37-0.62). Similarly, females aged 18 to 55 years with TTh had a lower risk of MACE (RR, 0.49; 95% CI, 0.28-0.85) and DVT (RR, 0.48; 95% CI, 0.25-0.93) and a similar risk of malignant breast neoplasm (RR, 0.62; 95% CI, 0.34-1.12). Females aged ≥56 years with TTh had a similar risk of MACE (RR, 0.84; 95% CI, 0.64-1.10), DVT (RR, 0.82; 95% CI, 0.50-1.36), and PE (RR, 0.52; 95% CI, 0.26-1.05) and a significantly lower risk of malignant breast neoplasm (RR, 0.51; 95% CI, 0.38-0.68). Risk of hirsutism was consistently higher in those with TTh as compared with propensity-matched controls. CLINICAL IMPLICATIONS: Our results contribute to safety data on TTh, a therapy for sexual dysfunction in women. STRENGTHS AND LIMITATIONS: The TriNetX Diamond Network allows for significant generalizability but has insufficient information for some factors. CONCLUSIONS: We found a decreased risk of MACE among women with TTh as compared with matched controls and a similar risk of MACE in postmenopausal women while demonstrating a similar or significantly lower risk of breast cancer on age-based subanalysis.

Perioperative Plasma in Addition to Red Blood Cell Transfusions Are Associated With Increased Venous Thromboembolism Risk Postoperatively.

BACKGROUND: Perioperative red blood cell (RBC) transfusions increase venous thromboembolic (VTE) events. Although a previous study found that plasma resuscitation after trauma was associated with increased VTE, the risk associated with additional perioperative plasma is unknown. METHODS: A US claims and EHR database (TriNetX Diamond Network) was queried. We compared surgical patients who received perioperative plasma and RBC to patients who received perioperative RBC but not plasma. Subanalyses included (1) all surgeries (n = 48,580) and (2) cardiovascular surgeries (n = 38,918). Propensity score matching was performed for age at surgery, ethnicity, race, sex, overweight and obesity, type 2 diabetes, disorders of lipoprotein metabolism, essential hypertension, neoplasms, nicotine dependence, coagulopathies, sepsis, chronic kidney disease, liver disease, nonsteroidal anti-inflammatory analgesics, platelet aggregation inhibitors, anticoagulants, hemoglobin level, outpatient service utilization, and inpatient services; surgery type was included for "all surgeries" analyses. Outcomes included 30-day mortality, postoperative VTE, pulmonary embolism (PE), and disseminated intravascular coagulation (DIC). RESULTS: After matching the surgical cohorts, compared to only RBC, plasma + RBC was associated with higher risk of postoperative mortality (4.52% vs 3.32%, risk ratio [RR]: 1.36 [95% confidence interval, 1.24-1.49]), VTE (3.92% vs 2.70%, RR: 1.36 [1.24-1.49]), PE (1.94% vs 1.33%, RR: 1.46 [1.26-1.68]), and DIC (0.96% vs 0.35%, RR: 2.75 [2.15-3.53]). Among perioperative cardiovascular patients, adding plasma to RBC transfusion was associated with similar increased risk. CONCLUSIONS: When compared with perioperative RBC transfusion, adding plasma was associated with increased 30-day postoperative mortality, VTE, PE, and DIC risk among surgical and cardiovascular surgical patients. Reducing unnecessary plasma transfusion should be a focus of patient blood management to improve overall value in health care.

Real-world comorbidities of atopic dermatitis in the US adult ambulatory population.

BACKGROUND: Atopic dermatitis (AD) is a pruritic, inflammatory skin disease associated with various comorbidities. However, comprehensive analyses of real-world comorbidities in adult patients with AD are limited. OBJECTIVE: To characterize the real-world comorbidities associated with adult AD in an ambulatory population. METHODS: We queried the MarketScan Commercial Claims and Encounters database from January 1, 2017 to December 31, 2017. Multivariable logistic regressions were performed to compare comorbidities in adult patients with AD versus age- and sex-matched controls. RESULTS: A total of 39,779 patients with AD and 353,743 controls were identified. Increased odds of psychiatric conditions, including anxiety (odds ratio [OR] 1.44) and mood disorders (OR 1.31), were observed in patients with AD. Patients with AD had higher likelihoods of autoimmune diseases, including vitiligo (OR 4.44) and alopecia areata (OR 6.01). Adult AD was also associated with infections, including impetigo (OR 9.72), methicillin-resistant Staphylococcus aureus (OR 3.92), and cellulitis (OR 2.52). Patients with AD were more likely to have systemic conditions, including lymphoid/hematopoietic malignancy (OR 1.91), atherosclerosis (OR 1.69), and metabolic syndrome (OR 1.47). For all the above, P < .001. LIMITATIONS: Retrospective analysis of health care claims data. CONCLUSION: Adult AD is associated with various psychiatric and systemic comorbidities, emphasizing the systemic nature of AD and the need for the collaborative management of these patients.

Clinical Practice Patterns for Surgical Shunts and Penile Prosthesis Placement in Men With Priapism: A Retrospective Large Claims Database Analysis.

INTRODUCTION: The objective of this study was to assess the rates of surgical shunting and prosthesis placement for acute ischemic priapism using a large multi-institutional claims database. METHODS: A US claims database network (TriNetX Diamond Network) was queried from 2010 to 2020. We constructed a cohort of men ages ≥ 16 years who (1) had a diagnosis of priapism and (2) underwent an irrigation of the corpora cavernosa for priapism. We assessed the number of men who then had a surgical penile shunt or penile prosthesis placement. Demographics, time to surgical procedure, and order of procedures were collected. RESULTS: A total of 6392 men were identified with the diagnosis of priapism and the procedure of corpora cavernosal irrigation. Of these men, 693 (11%) proceeded to surgical shunt. One hundred forty-four men (2%) underwent initial penile prosthesis placement. Of the men undergoing initial penile prosthesis, only 17 of 144 (12%) cases occurred within the first month of corpora cavernosal irrigation. Finally, when assessing choice of initial shunts vs initial penile prosthesis before and after 2015, overall rates of initial shunt (10.0% vs 8.5%, P < .0001) and initial prosthesis (3.1% vs 2.1%, P < .0001) were lower after 2015 when compared with rates prior to 2015. CONCLUSIONS: In this US claims-based analysis of men presenting with ischemic priapism and treated with initial irrigation, a small percentage (11%) of men went on to receive surgical shunting, and only 2% received an initial prosthesis. Men receiving initial prostheses were more likely to have more comorbidities, and overall surgical management of priapism has decreased over time.

Comparing risk of post infection erectile dysfunction following SARS Coronavirus 2 stratified by acute and long COVID, hospitalization status, and vasopressor administration: a U.S. large claims database analysis.

No study has yet assessed the risk of developing erectile dysfunction (ED) after a diagnosis of long COVID, defined by the Centers for Disease Control and Prevention as the persistence or presence of new symptoms at least 4 weeks after initial SARS-CoV-2 infection, when compared to those diagnosed with acute COVID or cases in which more severe treatment is required. To assess these risks, we queried the TriNetX COVID-19 Research Network from December 1st 2020 through June 2023. Men aged ≥ 18 diagnosed with long COVID were compared to those diagnosed with acute COVID and analyses were performed to compare men who were/were not hospitalized within 1 month of acute COVID diagnosis and men who did/did not need vasopressors. Cohorts were propensity score matched and compared for differences in new ED diagnosis and/or prescription of phosphodiesterase-5 inhibitors (PDE5i). After propensity score matching, the long and acute COVID cohorts included 2839 men with an average age of 54.5±16.7 and 55.1±17.1 years respectively (p = 0.21). Men with long COVID were more likely to develop ED or be prescribed PDE5i (3.63%) when compared to men with only acute COVID infections (2.61%) [RR 1.39; 95% CI 1.04, 1.87]. There was no statistically significant risk of developing ED or being prescribed PDE5i for individuals who received vasopressors [RR 0.92; 95% CI 0.77,1.10] or were hospitalized [RR 0.93; 95% CI 0.82,1.06].

Perioperative Pain Management With Opioid Analgesics in Colpopexy Increases Risk of New Persistent Opioid Usage.

IMPORTANCE: Although the use of perioperative pain medications is highly investigated, limited studies have examined the usage of pain medication for post hysterectomy prolapse repair and the few that have have been restricted to smaller sample sizes. OBJECTIVE: Our objective was to assess the association of perioperative opioid usage after posthysterectomy prolapse repairs with development of new persistent opioid usage. STUDY DESIGN: The TriNetX Diamond Research Network was queried to create our cohorts of opioid-naive adult women with vaginal repair or laparoscopic sacrocolpopexy. The primary study outcomes were (1) the rate of perioperative opioid usage and (2) development of new persistent opioid usage. All cohorts were matched on age, race, ethnicity, chronic kidney disease, hypertensive diseases, ischemic heart disease, diseases of the liver, obstructive sleep apnea, affective mood disorders, pelvic and perineal pain, obesity, tobacco use, and utilization of office/outpatient, inpatient, or emergency department services. RESULTS: We identified 10,414 opioid-naive women who underwent laparoscopic sacrocolpopexy and 13,305 opioid-naive women who underwent vaginal reconstruction. Rates of perioperative opioid usage were higher after laparoscopic sacrocolpopexy. Rates of developing new opioid usage were higher in both surgical-approach populations that received perioperative opioids compared with those that did not. Rates of new and persistent opioid usage did not differ by surgical approach when stratified by perioperative opioid usage. CONCLUSIONS: We identified that opioid dependence may occur after surgery if patients are given opioids within 7 days of either approach, associating opioid dependence with perioperative opioid usage rather than the approach taken.

Post-infection erectile dysfunction risk - comparing COVID-19 with other common acute viral infections: a large national claims database analysis.

It is unknown if the risk of erectile dysfunction (ED) following Coronavirus-19 (COVID-19) infection is virus-specific. Our study assessed the risk of ED in COVID-19 patients as compared to patients with other common viral infections. The TriNetX COVID-19 Research Network was queried. We examined cohorts of men aged ≥18 years infected with: COVID-19, influenza, respiratory syncytial virus, enterovirus, acute viral hepatitis, mononucleosis, and herpes zoster. Men were included if they had at least one outpatient follow-up visit within 18 months and excluded if they had one of the other viruses of interest or a prior ED diagnosis or treatment, prostatectomy, pelvis radiation, or chronic hepatitis infection. Cohorts were propensity score matched and compared for differences in new ED diagnosis and/or prescription of phosphodiesterase-5 inhibitors (PDE5i). COVID-19 positive men were less likely to develop ED or have a PDE5i prescription than men with infected with herpes zoster [Relative Risk (RR): 0.37, 95% Confidence Interval (CI) 0.27-0.49] and more likely to develop ED or have a PDE5i prescription than men with no acute viral illness (RR: 1.33, 95% CI 1.25-1.42). In this national propensity-matched cohort study comparing post-infection ED risk and PDE5i prescriptions, we found that COVID-19 was no more likely to result in a diagnosis of ED or prescription of PDE5i when compared to all acute viral illnesses except herpes zoster, which was more likely to result in a diagnosis of ED or prescription of PDE5i when compared to COVID-19. These findings suggest an inflammatory etiology (perhaps due to cytokine release, endothelial dysfunction, or blunted hormone signaling) behind any acute infection can result in a heightened ED risk; however, further studies are required to investigate the connection between other viral infections and ED.

Involvement of vulva in lichen sclerosus increases the risk of antidepressant and benzodiazepine prescriptions for psychiatric disorder diagnoses.

While vulvar lichen sclerosus (VLS) causes intense pruritus, associated risks of mood disorders and prescription patterns and impact of concurrent sexual dysfunction are unknown. We queried TriNetX Diamond Network between 2009 and 2022, conducting three comparisons after propensity-score matching for demographics and relevant comorbidities: (1) women with lichen sclerosus (LS) sparing the vulva vs. women with VLS; (2) VLS patients who received treatment within 6 months of diagnosis vs. patients who did not and (3) VLS patients with vs. without sexual dysfunction. Outcomes included new depressive episodes, anxiety disorder, major depressive disorder (MDD), and prescriptions of antidepressants or benzodiazepines. After matching, VLS was associated with increased depressive episode [risk ratio (RR) 1.39], anxiety disorder (RR 1.93), and MDD (RR 2.00) diagnoses compared to LS sparing the vulva. Next, VLS treatment was associated with decreased risk of depressive episode (RR 0.60) and anxiety disorder (RR 0.72). Finally, concurrent sexual dysfunction was associated with increased benzodiazepine (RR 3.50), vaginal estrogen (RR 6.20), antipruritic agents (RR 3.90), and topical anti-inflammatory (RR 2.61) prescriptions. In conclusion, vulvar involvement is associated with increased risk of antidepressant and benzodiazepine prescriptions, and diagnosis of depressive episode, anxiety disorder, or MDD.

Single-strand mismatch and damage patterns revealed by single-molecule DNA sequencing.

Mutations accumulate in the genome of every cell of the body throughout life, causing cancer and other genetic diseases1-4. Almost all of these mosaic mutations begin as nucleotide mismatches or damage in only one of the two strands of the DNA prior to becoming double-strand mutations if unrepaired or misrepaired5. However, current DNA sequencing technologies cannot resolve these initial single-strand events. Here, we developed a single-molecule, long-read sequencing method that achieves single-molecule fidelity for single-base substitutions when present in either one or both strands of the DNA. It also detects single-strand cytosine deamination events, a common type of DNA damage. We profiled 110 samples from diverse tissues, including from individuals with cancer-predisposition syndromes, and define the first single-strand mismatch and damage signatures. We find correspondences between these single-strand signatures and known double-strand mutational signatures, which resolves the identity of the initiating lesions. Tumors deficient in both mismatch repair and replicative polymerase proofreading show distinct single-strand mismatch patterns compared to samples deficient in only polymerase proofreading. In the mitochondrial genome, our findings support a mutagenic mechanism occurring primarily during replication. Since the double-strand DNA mutations interrogated by prior studies are only the endpoint of the mutation process, our approach to detect the initiating single-strand events at single-molecule resolution will enable new studies of how mutations arise in a variety of contexts, especially in cancer and aging.

Malic enzyme 2 connects the Krebs cycle intermediate fumarate to mitochondrial biogenesis.

Mitochondria have an independent genome (mtDNA) and protein synthesis machinery that coordinately activate for mitochondrial generation. Here, we report that the Krebs cycle intermediate fumarate links metabolism to mitobiogenesis through binding to malic enzyme 2 (ME2). Mechanistically, fumarate binds ME2 with two complementary consequences. First, promoting the formation of ME2 dimers, which activate deoxyuridine 5'-triphosphate nucleotidohydrolase (DUT). DUT fosters thymidine generation and an increase of mtDNA. Second, fumarate-induced ME2 dimers abrogate ME2 monomer binding to mitochondrial ribosome protein L45, freeing it for mitoribosome assembly and mtDNA-encoded protein production. Methylation of the ME2-fumarate binding site by protein arginine methyltransferase-1 inhibits fumarate signaling to constrain mitobiogenesis. Notably, acute myeloid leukemia is highly dependent on mitochondrial function and is sensitive to targeting of the fumarate-ME2 axis. Therefore, mitobiogenesis can be manipulated in normal and malignant cells through ME2, an unanticipated governor of mitochondrial biomass production that senses nutrient availability through fumarate.