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Processing bodies (P-bodies) are cytoplasmic membrane-less organelles which host multiple mRNA processing events. While the fundamental principles of P-body organization are beginning to be elucidated in vitro , a nuanced understanding of how their assembly is regulated in vivo remains elusive. Here, we investigate the potential link between ER exit sites and P-bodies in Drosophila melanogaster egg chambers. Employing a combination of live and super-resolution imaging, we found that P-bodies associated with ER exit sites are larger and less mobile than cytoplasmic P-bodies, indicating that they constitute a distinct class of P-bodies which are more mature than their cytoplasmic counterparts. Moreover, we demonstrate that altering the composition of ER exit sites has differential effects on core P-body proteins (Me31B, Cup, and Trailer Hitch) suggesting a potential role for ER exit sites in P-body organization. We further show that in the absence of ER exit sites, P-body integrity is compromised and the stability and translational repression efficiency of the maternal mRNA, oskar , are reduced. Finally, we show that ER stress is communicated to P-bodies via ER exit sites, highlighting the pivotal role of ER exit sites as a bridge between membrane-bound and membrane-less organelles in ER stress response. Together, our data unveils the significance of ER exit sites not only in governing P-body organization, but also in facilitating inter-organellar communication during stress, potentially bearing implications for a variety of disease pathologies., Competing Interests: DISCLOSURE STATEMENT There are no potential conflicts of interests.

Introduction: The results of the phase 3 ALSYMPCA trial showed that Radium-223 (Ra-223) improves overall survival (OS) and delays onset of first symptomatic skeletal event vs. placebo in patients with metastatic castration-resistant prostate cancer (mCRPC). The purpose of the REACTIVATE study was to inform the optimal placement of Ra-233 in the treatment sequence by evaluating clinical outcomes and healthcare resource utilization using real-world data from multiple Canadian provinces., Methods: This retrospective cohort study analyzed patient outcomes according to Ra-223 placement using administrative databases of four Canadian provinces, encompassing 4301 patients with mCRPC who received at least two lines of life-prolonging therapy (LPT) for mCRPC. Outcomes included OS, event-free survival (EFS), and healthcare resource utilization. Each province was analyzed separately., Results: OS, measured from the start of second-line LPT, differed between provinces: those in Ontario receiving second-line Ra-223 had a longer OS vs. those receiving it in third-line or later (hazard ratio [HR ] 0.79, 95% confidence interval [CI] 0.66-0.95). There was no difference between lines of therapy in patients in British Columbia (HR 1.165, 95% CI, 0.894-1.518, p=0.2576), and OS was numerically worse but not statistically significant in patients receiving Ra-223 in second-line in Quebec (HR 1.44, 95% CI, 0.93-2.24). Other outcomes also varied across provinces, with second-line use of Ra-223 being associated with longer EFS and reduced healthcare utilization vs. third-line use in Ontario but not in Quebec., Conclusions: Significant heterogeneity exists in the management and outcomes of mCRPC between provinces, particularly regarding the placement of Ra-223 in the treatment sequence.

Introduction: Bone-targeted therapies (BTTs) are integral to the management of bone metastases in metastatic castration-resistant prostate cancer (mCRPC). BTTs vary considerably in referral and drug access pathways and optimal BTT use requires multi-specialty consultation and supervision. Health quality improvement (HQI) has become the predominant framework to improve patient care in multidisciplinary settings., Methods: HQI initiatives on use of BTT in mCRPC were developed and evaluated in five centers of a provincial cancer center network using Plan-Do-Study-Act (PDSA) methodology. Multidisciplinary teams (MDTs) completed a common quality assessment form and an HQI template and then implemented an HQI initiative. Feedback and findings were shared and discussed at regional events. It was subsequently determined whether to adopt, adapt, or abandon initiatives., Results: Patterns of unmet needs varied across type of BTT. Gaps in use of radium-223 were mostly referral and education issues that could be directly addressed at the local level by participating clinician teams. Conversely, most supportive BTT gaps were related to coverage and resourcing support. HQI initiatives selected by each site consisted of implementation or expansion of local MDT meetings, referral documents, databases, and improvement charters. The main HQI initiative was completed in four sites and was adapted or adopted in three. Improvements in BTT use were observed in two of three centers with data on HQI process measures., Conclusions: Despite the overall heterogenous structure of the groups and metrics used, this study demonstrated that the PDSA framework provides the needed structure for improvements in BTT use in mCRPC across multiple sites.

Objective: As vulvar and vaginal cancers are rare malignancies, treatment is extrapolated from the cervical cancer field. Further studies are necessary to evaluate whether surgery, radiotherapy (RT), or combined chemoRT is most beneficial., Methods: A retrospective chart review was conducted on patients diagnosed with vulvar or vaginal cancer in 2000-2017. Descriptive statistics was used to summarize demographic factors. Kaplan-Meier curves, log-rank tests, multivariate analysis with hazard ratios (HR) were conducted to compare survival outcomes, including overall survival (OS), disease-free survival, and cancer-specific survival, between surgery, RT, and chemoRT., Results: This study included 688 patients with either vulvar (n = 560, 81%) or vaginal cancer (n = 128, 19%). Median age of diagnosis was 68 (27-98) years. In multivariate survival analysis, vulvar cancer was associated with more likelihood of death (HR: 1.50, p = 0.042) compared to vaginal cancer. For patients who received definitive RT, median OS was 63.8 months with concurrent chemotherapy vs. 46.3 months without for vulvar cancer (p = 0.75); for vaginal, median OS 100.4 with chemotherapy vs. 66.6 months without (p = 0.31). For vulvar cancer patients who received RT (n = 224), adding chemotherapy (n = 100) was not associated with statistically significant OS improvement (HR: 0.989, p = 0.957). Similarly, vaginal cancer patients who received chemoRT (n = 51) did not have significant OS benefit (HR: 0.720, p = 0.331) over patients who received RT (n = 49)., Conclusions: In this retrospective study, chemoRT was not associated with significant improvements in survival compared to RT in vulvar or vaginal cancer. Future studies investigating novel therapies to treat these cancers are needed to improve patient outcomes., Competing Interests: Declaration of Competing Interest All authors declare no conflicts of interest pertaining to this work., (Copyright © 2023 Elsevier Inc. All rights reserved.)

The PACIFIC trial showed a survival benefit with durvalumab through five years in stage III unresectable non-small cell lung cancer (NSCLC). However, optimal use of imaging to detect disease progression remains unclearly defined for this population. An expert working group convened to consider available evidence and clinical experience and develop recommendations for follow-up imaging after concurrent chemotherapy and radiation therapy (CRT). Voting on agreement was conducted anonymously via online survey. Follow-up imaging was recommended for all suitable patients after CRT completion regardless of whether durvalumab is received. Imaging should occur every 3 months in Year 1, at least every 6 months in Year 2, and at least every 12 months in Years 3-5. Contrast computed tomography was preferred; routine brain imaging was not recommended for asymptomatic patients. The medical oncologist should follow-up during Year 1 of durvalumab therapy, with radiation oncologist involvement if pneumonitis is suspected; medical and radiation oncologists can subsequently alternate follow-up. Some patients can transition to the family physician/community primary care team at the end of Year 2. In Years 1-5, patients should receive information regarding smoking cessation, comorbidity management, vaccinations, and general follow-up care. These recommendations provide guidance on follow-up imaging for patients with stage III unresectable NSCLC whether or not they receive durvalumab consolidation therapy.

Background: Factors that impact recurrence in stages IB to IV include larger tumor, high-risk histology, older age, and lymphovascular invasion (LVI); however, local studies on risk factors for recurrence in British Columbia and our local recurrence patterns have not been well studied. Furthermore, the efficacy of treatment modalities including surgery and chemoradiation in the different stages of cervical cancer have not been clarified in this population., Objectives: The purpose of this study is to determine the disease and treatment characteristics of stages IB to IV cervical cancer which are associated with survival differences within British Columbia., Methods/design: We performed a retrospective population study. A chart review on cervical cancer patients in British Columbia between 1 January 2010 and 31 December 2017 was done. Demographic data and treatment details were collected. Data were analyzed using multivariate Cox regressions, pairwise comparison using the Log-Rank test, and chi-square tests., Results: We included 780 patients (stage I: 31.5%, II: 20.0%, III: 34.5%, and IV: 3.3%). LVI and p16 negativity were associated with decreased overall survival (OS), and multivariate analyses show them to be independent risk factors for poorer survival. Surgical resection in stage I was associated with improved survival, but not with stages II-IV. The use of radical radiation therapy (RT), brachytherapy, and concurrent chemotherapy were independently associated with improved survival in stages II-IV. Peri-RT chemotherapy was not associated with survival benefit in adeno/adenosquamous carcinoma. There were 180 recurrences (23.1%), mostly distant metastases (42.8%). There were fewer recurrences after resection of tumors <2 cm compared to tumors 2 cm or larger (6.49% vs 31.3%, p = 0.00011). Only 37.7% of recurrence/metastases were treated with first-line carboplatin/paclitaxel/bevacizumab, but it was associated with better OS compared to other regimens (median OS 40.1 vs 24.8 months, p = 0.03)., Conclusion: A significant portion of patients with localized cervical cancer relapse despite radical therapy, with LVI and p16 negativity associated with poorer survival. Surgical resection may still play a role in stage IB disease, while RT, brachytherapy, and concurrent chemotherapy should be considered first-line therapy in stage II-IV diseases. First-line carboplatin, paclitaxel, and bevacizumab for recurrence shows improved survival.

Non-obstructive azoospermia is a major clinical issue associated with male infertility that remains to be addressed. Although neogenin is reportedly abundantly expressed in the testis, its role in mammalian spermatogenesis is unknown. We systematically investigated the role of neogenin during spermatogenesis by performing loss-of-function studies. Testis-specific neogenin conditional knock-out (cKO) mice were generated using CRISPR/Cas9 and neogenin-targeting guide RNAs. We analyzed the expression profiles of germ cell factors by RT-PCR and Western blotting. Neogenin localized mainly to spermatogonia in seminiferous tubules of mouse testes. RT-PCR and Western blot analyses further demonstrated that neogenin expression varied during spermatogenesis and was dramatically increased at postnatal day 12-25 during the pubertal stage. In neogenin-cKO mouse testes, the ratio of primary and secondary spermatocytes was significantly decreased compared with the control, while the number of apoptotic testicular cells was significantly increased. Taken together, these results suggest that neogenin plays a pivotal role in the maintenance and proliferation of spermatogonia during the early stage of spermatogenesis in mice.

Introduction: Unresectable and metastatic small cell carcinoma of the prostate (SCPC) is a rare and aggressive disease that is under-represented in clinical trials. We carried out a retrospective chart review of metastatic or unresectable SCPC patients at British Columbia (BC) Cancer centers, studying diagnosis and treatment patterns., Methods: Drug-dispensing records from the six BC Cancer centers were obtained from 2002-2017. For each patient, information was collected on baseline information prior to therapy and for each line of treatment. Treatments at each line were compared regarding time to progression and overall survival by Kaplan-Meier curves., Results: Forty-one patients received treatment; 65.6% had metastatic disease and 61% had pure small cell carcinoma. Median time from treatment to death was 10 months (95% confidence interval [CI] 6-16). Patients with initially prostate-confined disease had a better median overall survival (mOS) of 21 months (95% CI 13-34) compared to those with initially locally advanced (mOS 19 months, 95% CI 5-37) and metastatic disease (mOS 8 months, 95% CI 6-10) (log-rank p=0.0364). All patients received either cisplatin- or carboplatin-based combination chemotherapy as the first-line treatment and 36.7% received second-line therapy. Time to second-line therapy was eight months for those who presented with metastatic SCPC, compared to 13 months for those with initial non-metastatic SCPC., Conclusions: This single-province, multi-institution cohort reports data on unresectable and metastatic SCPC and highlights the poor prognosis of this rare disease entity.

Acute hepatic porphyria (AHP) is a group of rare genetic diseases of heme biosynthesis resulting in severe neurovisceral attacks and chronic complications that negatively impact patients' well-being. This study evaluated the impacts of AHP on patients' physical and emotional health from a global perspective. Adult patients from the United States, Italy, Spain, Australia, Mexico, and Brazil with AHP with >1 porphyria attack within the past 2 years or receiving intravenous hemin and/or glucose for attack prevention completed an online survey assessing demographics, health characteristics, and patient-reported outcomes. Results were analyzed collectively and by patient subgroups. Ninety-two patients with AHP across the six countries completed the survey. More than 70% of patients reported that their physical, emotional, and financial health was fair or poor. Among patients who reported pain, fatigue, and muscle weakness, 94.3%, 95.6%, and 91.4%, respectively, reported that these symptoms limited daily activities. Moderate to severe depression was present in 58.7% of patients, and moderate to severe anxiety in 48.9% of patients. Of the 47% of patients who were employed, 36.8% reported loss in productivity while at work. Among patients, 85.9% reported that they had to change or modify goals that were important to them because of AHP. Aside from differences in healthcare utilization and pain severity, scores did not significantly vary with attack rate or use of hemin or glucose prophylactic treatments. AHP substantially impacts patients' physical and emotional well-being, regardless of hemin or glucose prophylactic treatment or frequency of attacks. This multinational study demonstrates that there is substantial disease burden for patients with AHP, even among those experiencing sporadic attacks or using prophylactic treatment., Competing Interests: The authors declare the following competing interests: Amy Dickey had speaking engagements and received consulting honoraria from Alnylam Pharmaceuticals for participation in this research and for other porphyria‐related consulting. Kristen Wheeden was employed by the American Porphyria Foundation at the time of the study and currently is president of the United Porphyrias Association; she received grant and sponsorship funding to the American Porphyria Foundation from Alnylam Pharmaceuticals and served on a medical advisory board for Alnylam Pharmaceuticals. Desiree Lyon received grant and sponsorship funding to the American Porphyria Foundation from Alnylam Pharmaceuticals. Sue Burrell received grant and sponsorship funding to the British Porphyria Association as well as consulting honoraria from Alnylam Pharmaceuticals for participation on various patient advisory group leadership advisory boards. Sean Hegarty, Rocco Falchetto, and Jasmin Barman‐Aksözen report no competing interests. Edrin R. Williams was employed by the American Porphyria Foundation at the time of the study. Marc DeCongelio, Alison Bulkley, Joana E. Matos, and Tarek Mnif were employed at the time of the study by Kantar Health (now Cerner Enviza), a company that provides consultancy support to Alnylam. Jordanna Mora and John J. Ko were employed by Alnylam Pharmaceuticals at the time of the study and own stock and stock options in the company; currently they are employed by Beam Therapeutics and own stock and stock options in the company. Stephen Meninger and Stephen Lombardelli are employed by and own stock and stock options in Alnylam Pharmaceuticals. Danielle Nance served on advisory boards for Aptevo Therapeutics, Bayer, HemaBiologics, and Medexus Pharmaceuticals; served on speaker bureaus for BPL and the National Hemophilia Foundation; provided consulting services to Goval; served as an author for Bayer; and had speaking engagements and received consulting honoraria from Alnylam Pharmaceuticals for participation in this research., (© 2022 The Authors. JIMD Reports published by John Wiley & Sons Ltd on behalf of SSIEM.)

Introduction: Acute hepatic porphyria (AHP) is a family of rare metabolic diseases characterized by potentially life-threatening acute attacks and, in some patients, chronic debilitating symptoms. While patients with frequent or recurrent attacks (three or more attacks annually) are known to have reduced health-related quality of life (HRQoL) as most aspects of daily living are impacted, limited data exist in patients with sporadic attacks. This research aims to identify porphyria-related symptoms between attacks, characterize the frequency, severity, and bothersomeness of these symptoms, and more generally understand the burden of this disease in patients who experience attacks sporadically., Methods: Patients with AHP with sporadic attacks (AHP-SA) (at least one porphyria attack in the past 2 years, but no more than two attacks per year in the previous 2 years) were recruited, via outreach performed by patient advocacy groups, for participation in qualitative telephone interviews. Interviews were conducted using a semi-structured guide and were audio-recorded, transcribed, anonymized, coded, and analyzed to determine if saturation was reached., Results: A total of 14 participants with AHP-SA were interviewed (mean age 45 years, 100% female). The most frequently reported chronic symptoms were fatigue, pain, heartburn, and constipation. The most frequently experienced chronic impacts were difficulty performing daily activities, difficulty exercising, negative impact on work, need for a special diet, anxiety, and depression. Beyond these chronic symptoms and impacts, participants also frequently described flares in their porphyria that were severe, did not qualify in their minds as an acute attack, but were nonetheless more severe than their typical chronic experience., Conclusion: Patients with acute hepatic porphyria who experience sporadic attacks face significant chronic symptoms and impacts that frequently require significant pharmacological and clinical treatment. The reported severity of these symptoms and impacts suggests that the humanistic burden of AHP-SA is substantial and may lead to a significant decrease in health-related quality of life in these patients between acute attacks. The presence of flares that do not reach the level of what is considered an acute attack by patients is a unique finding of this study not reported elsewhere and requires additional investigation., (© 2022. The Author(s).)

Objectives: Patient-derived induced pluripotent stem cells (iPSCs) are materials that can be used for autologous stem cell therapy. We screened mtDNA mutations in iPSCs and iPSC-derived neuronal cells from patients with Alzheimer's disease (AD). Also, we investigated whether the mutations could affect mitochondrial function and deposition of β-amyloid (Aβ) in differentiated neuronal cells., Materials and Methods: mtDNA mutations were measured and compared among iPSCs and iPSC-derived neuronal cells. The selected iPSCs carrying mtDNA mutations were subcloned, and then their growth rate and neuronal differentiation pattern were analyzed. The differentiated cells were measured for mitochondrial respiration and membrane potential, as well as deposition of Aβ., Results: Most iPSCs from subjects with AD harbored ≥1 mtDNA mutations, and the number of mutations was significantly higher than that from umbilical cord blood. About 35% and 40% of mutations in iPSCs were shared with isogenic iPSCs and their differentiated neuronal precursor cells, respectively, with similar or different heteroplasmy. Furthermore, the mutations in clonal iPSCs were stable during extended culture and neuronal differentiation. Finally, mtDNA mutations could induce a growth advantage with higher viability and proliferation, lower mitochondrial respiration and membrane potential, as well as increased Aβ deposition., Conclusion: This study demonstrates that mtDNA mutations in patients with AD could lead to mitochondrial dysfunction and accelerated Aβ deposition. Therefore, early screening for mtDNA mutations in iPSC lines would be essential for developing autologous cell therapy or drug screening for patients with AD., (© 2022 The Authors. Cell Proliferation published by European Cell Proliferation Society and John Wiley & Sons Ltd.)

Millions of people around the world suffer from infertility, with the number of infertile couples and individuals increasing every year. Assisted reproductive technologies (ART) have been widely developed in recent years; however, some patients are unable to benefit from these technologies due to their lack of functional germ cells. Therefore, the development of alternative methods seems necessary. One of these methods is to create artificial oocytes. Oocytes can be generated in vitro from the ovary, fetal gonad, germline stem cells (GSCs), ovarian stem cells, or pluripotent stem cells (PSCs). This approach has raised new hopes in both basic research and medical applications. In this article, we looked at the principle of oocyte development, the landmark studies that enhanced our understanding of the cellular and molecular mechanisms that govern oogenesis in vivo, as well as the mechanisms underlying in vitro generation of functional oocytes from different sources of mouse and human stem cells. In addition, we introduced next-generation ART using somatic cells with artificial oocytes. Finally, we provided an overview of the reproductive application of in vitro oogenesis and its use in human fertility.

Background & Aims: Upregulation of hepatic delta-aminolevulinic acid synthase 1 with accumulation of potentially toxic heme precursors delta-aminolevulinic acid and porphobilinogen is fundamental to the pathogenesis of acute hepatic porphyria., Aims: evaluate long-term efficacy and safety of givosiran in acute hepatic porphyria., Methods: Interim analysis of ongoing ENVISION study (NCT03338816), after all active patients completed their Month 24 visit. Patients with acute hepatic porphyria (≥12 years) with recurrent attacks received givosiran (2.5 mg/kg monthly) (n = 48) or placebo (n = 46) for 6 months (double-blind period); 93 received givosiran (2.5 mg or 1.25 mg/kg monthly) in the open-label extension (continuous givosiran, n = 47/48; placebo crossover, n = 46/46). Endpoints included annualized attack rate, urinary delta-aminolevulinic acid and porphobilinogen levels, hemin use, daily worst pain, quality of life, and adverse events., Results: Patients receiving continuous givosiran had sustained annualized attack rate reduction (median 1.0 in double-blind period, 0.0 in open-label extension); in placebo crossover patients, median annualized attack rate decreased from 10.7 to 1.4. Median annualized days of hemin use were 0.0 (double-blind period) and 0.0 (open-label extension) for continuous givosiran patients and reduced from 14.98 to 0.71 for placebo crossover patients. Long-term givosiran led to sustained lowering of delta-aminolevulinic acid and porphobilinogen and improvements in daily worst pain and quality of life. Safety findings were consistent with the double-blind period., Conclusions: Long-term givosiran has an acceptable safety profile and significantly benefits acute hepatic porphyria patients with recurrent attacks by reducing attack frequency, hemin use, and severity of daily worst pain while improving quality of life., (© 2021 The Authors. Liver International published by John Wiley & Sons Ltd.)

When ejaculated sperm travels through the vagina to the uterus, mucus secreted by the cervical canal generally filters out sperm having low motility and poor morphology. To investigate this selection principle in vivo, we developed a microfluidic sperm-sorting chip with a viscous medium (polyvinylpyrrolidone: PVP) to imitate the biophysical environment mimic system of the human cervical canal. The material property of the PVP solution was tuned to the range of viscosities of cervical mucus using micro-viscometry. The selection of high-quality human sperm was experimentally evaluated in vitro and theoretically analyzed by the convection-diffusion mechanism. The convection flow is shown to be dominant at low viscosity of the medium used in the sperm-sorting chip when seeded with raw semen; hence, the raw semen containing sperm and debris convectively flow together with suppressed relative dispersions. Also, it was observed that the sperm selected via the chip not only had high motilities but also normal morphologies and high DNA integrity. Therefore, the biomimetic sperm-sorting chip with PVP medium is expected to improve male fertility by enabling the selection of high-quality sperm as well as uncovering pathways and regulatory mechanisms involved in sperm transport through the female reproductive tract for egg fertilization.

Background: Real-world data on palliative systemic therapies (PST) in treating metastatic bladder cancer (mBC) is limited. This study investigates current trends in treating mBC with first- (1L) and second-line (2L) chemotherapy (CT) and immunotherapy (IT)., Methods: A chart review was conducted on patients diagnosed with stage II-IV bladder cancer in 2014-2016. Survival outcomes were compared between chemotherapy, immunotherapy, and supportive care., Results: out of 297 patients, 77% were male. 44% had stage IV disease at diagnosis. Median age at metastasis was 73 years. 40% of patients received 1L PST and 34% received 2L PST. Median overall survival (mOS) was longer in those receiving PST versus no treatment ( p < 0.001). Patients receiving CT and IT sequentially had the longest mOS (18.99 months). First-line IT and CT mOS from treatment start dates were 5.03 and 9.13 months, respectively ( p = 0.81). Gemcitabine with cisplatin (8.88 months) or carboplatin (9.13 months) were the most utilized 1L chemotherapy regimens ( p = 0.85). 2L IT and CT mOS from treatment start dates were 6.72 and 3.78 months, respectively ( p = 0.15)., Conclusion: real-world mOS of >1.5 years in mBC is unprecedented and supports using multiple lines of PST. Furthermore, immunotherapy may be a comparable alternative to chemotherapy in both 1L and 2L settings.

The maturation of the oocyte is influenced by cumulus cells (CCs) and associated with pregnancy rate, whereas the influencing factors have not been completely elucidated in the CCs. In this study, we identified new regulators of CCs for high-quality oocytes and successful pregnancies during assisted reproductive techniques. CCs were collected from cumulus-oocyte complexes (COCs) in young (≤33 years old) and old (≥40 years old) women undergoing intracytoplasmic sperm injection (ICSI) procedures. We screened for factors differentially expressed between young vs. old CCs and pregnancy vs. non-pregnancy using whole mRNA-seq-next-generation sequencing (NGS). We characterized the transcriptome of the CCs to identify factors critical for achieving pregnancy in IVF cycles. Women in the young and old pregnancy groups exhibited the up- and downregulation of multiple genes compared with the non-pregnancy groups, revealing the differential regulation of several specific genes involved in ovarian steroidogenesis in CCs. It was shown that the low-density lipoprotein (LDL) receptor to the steroidogenesis pathway was upregulated in CCs with higher maturity rates of oocytes in the pregnancy group. In conclusion, a higher pregnancy rate is related to the signaling pathway of steroidogenesis by the LDL receptor in infertile women undergoing IVF procedures.

The purpose of this study was to investigate whether polymorphisms in five microRNAs (miRNAs), miR-604 A>G, miR-608 C>G, 631 I/D, miR-938 G>A, and miR-1302-3 C>T, are associated with the risk of idiopathic recurrent pregnancy loss (RPL). Blood samples were collected from 388 patients with idiopathic RPL (at least two consecutive spontaneous abortions) and 227 control participants. We found the miR-604 AG and AG + GG genotypes of miR-604 , the miR-938 GA and GA + AA genotypes of miR-938 , and the miR-1302-3 CT and CT + TT genotypes of miR-1302-3 are less frequent than the wild-type (WT) genotypes, miR-604 AA, miR-938 GG, and miR-1302-3 CC, respectively, in RPL patients. Using allele-combination multifactor dimensionality reduction (MDR) analysis, we found that eight haplotypes conferred by the miR-604/miR-608/miR-631/miR-938/miR-1302-3 allele combination, A-C-I-G-T, A-C-I-A-C, G-C-I-G-C, G-C-I-G-T, G-G-I-G-C, G-G-I-G-T, G-G-I-A-C, G-G-D-G-C, three from the miR-604/miR-631/miR-938/miR-1302-3 allele combination, A-I-G-T, G-I-G-C, G-I-A-T, one from the miR-604/miR-631/miR-1302-3 allele combination, G-I-C, and two from the miR-604/miR-1302-3 allele combination, G-C and G-T, were less frequent in RPL patients, suggesting protective effects (all p < 0.05). We also identified the miR-604 A>G and miR-938 G>A polymorphisms within the seed sequence of the mature miRNAs and aligned the seed sequences with the 3'UTR of putative target genes, methylenetetrahydrofolate reductase ( MTHFR ) and gonadotropin-releasing hormone receptor ( GnRHR ), respectively. We further found that the binding affinities between miR-604/miR-938 and the 3'UTR of their respective target genes ( MTHFR, GnRHR ) were significantly different for the common ( miR-604 A, miR-938 G) and variant alleles ( miR-604 G, miR-938 A). These results reveal a significant association between the miR-604 A>G and miR-938 G>A polymorphisms and idiopathic RPL and suggest that miRNAs can affect RPL in Korean women.

Objectives: This study retrospectively compares the effectiveness of methylprednisolone to dexamethasone in patients with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2 or COVID-19) requiring intensive care., Design: This is an institutional review board approved cohort study in patients with COVID-19 requiring intensive care unit (ICU) admission. Patients admitted and requiring oxygen supplementation were treated with no steroids, methylprednisolone, or dexamethasone., Setting: This study takes place in the ICU's at a large, tertiary, public teaching hospital serving a primarily low-income community in urban Los Angeles., Patients: All eligible patients admitted to the ICU for COVID-19 respiratory failure from March 1 to July 31, 2020 were included in this study., Interventions: A total of 262 patients were grouped as receiving usual care (n = 75), methylprednisolone dosed at least at 1mg/kg/day for ≥ 3 days (n = 104), or dexamethasone dosed at least at 6 mg for ≥7 days (n = 83)., Measurements and Main Results: All-cause mortality within 50 days of initial corticosteroid treatment as compared to usual care was calculated. The mortality effect was then stratified based on levels of respiratory support received by the patient. In this cohort of 262 patients with severe COVID-19, all-cause mortalities in the usual care, methylprednisolone, and dexamethasone groups were 41.3%, 16.4% and 26.5% at 50 days ( P < 0.01) respectively. In patients requiring mechanical ventilation, mortality was 42% lower in the methylprednisolone group than in the dexamethasone group (hazard ratio 0.48, 95% CI: 0.235-0.956, P = 0.0385)., Conclusions: In COVID-19 patients requiring mechanical ventilation, sufficiently dosed methylprednisolone can lead to a further decreased mortality as compared to dexamethasone.

Inflammation is a major cause of several chronic diseases and is reported to be recovered by the immuno-modulation of mesenchymal stem cells (MSCs). While most studies have focussed on the anti-inflammatory roles of MSCs in stem cell therapy, the impaired features of MSCs, such as the loss of homeostasis by systemic aging or pathologic conditions, remain incompletely understood. In this study, we investigated whether the altered phenotypes of human placenta-derived MSCs (hPD-MSCs) exposed to inflammatory cytokines, including TNF-α and IFN-γ, could be protected by MIT-001, a small anti-inflammatory and anti-necrotic molecule. MIT-001 promoted the spindle-like shape and cytoskeletal organization extending across the long cell axis, whereas hPD-MSCs exposed to TNF-α/IFN-γ exhibited increased morphological heterogeneity with an abnormal cell shape and cytoskeletal disorganization. Importantly, MIT-001 improved mitochondrial distribution across the cytoplasm. MIT-001 significantly reduced basal respiration, ATP production, and cellular ROS levels and augmented the spare respiratory capacity compared to TNF-α/IFN-γ-exposed hPD-MSCs, indicating enhanced mitochondrial quiescence and homeostasis. In conclusion, while TNF-α/IFN-γ-exposed MSCs lost homeostasis and mitochondrial quiescence by becoming over-activated in response to inflammatory cytokines, MIT-001 was able to rescue mitochondrial features and cellular phenotypes. Therefore, MIT-001 has therapeutic potential for clinical applications to treat mitochondrion-related inflammatory diseases.

Mesenchymal stem cells (MSCs) are multipotent cells with critical roles in homeostasis and regeneration. MSCs undergo aging in response to various stresses, and this causes many diseases including degenerative disorders. Thus, regulation of aging factors is crucial for healthy aging. Mitochondrial open reading frame of the 12S rRNA-c (MOTS-c) was recently reported to regulate metabolic homeostasis. Here, we investigated the restorative effects of MOTS-c on aged human placenta-derived MSCs (hPD-MSCs). MOTS-c promoted the morphology of old hPD-MSCs. MOTS-c significantly activated AMP-activated protein kinase, which is the main target pathway of MOTS-c, and inhibited its antagonistic effector mTORC1. MOTS-c considerably enhanced mitochondrial homeostasis by decreasing oxygen consumption and reactive oxygen species production. The mitochondrial state of MOTS-c-treated old hPD-MSCs was more similar to that of young hPD-MSCs than the mitochondrial state of non-treated old hPD-MSCs. MOTS-c also decreased lipid synthesis. In conclusion, we demonstrated that MOTS-c promotes homeostasis in aged hPD-MSCs., (Copyright © 2021 Elsevier B.V. and Mitochondria Research Society. All rights reserved. All rights reserved.)

Background: This study used quantitative and qualitative research methods to analyze how acute hepatic porphyria (AHP) affects patients with varying annualized porphyria attack rates. The overall impact of AHP on patients and caregivers, including their quality of life, was explored. The nature and treatment of acute attacks, experiences of long-term heme arginate treatment and access to other appropriate treatment, and the extent of and treatment for chronic symptoms were also investigated within this study., Methods: Patient and caregiver data were collected via an online survey of members of the British Porphyria Association, followed by an optional 1-h telephone interview., Results: Thirty-eight patients and 10 caregivers responded to the survey. Of those, 10 patients and three caregivers completed follow-up interviews. Overall, 19 patients (50%) had experienced an acute attack within the previous 2 years, and the severity and types of symptoms experienced during or between acute attacks varied considerably. There were no clear definitions among patients for 'mild' or 'severe' attacks. Treatments and treatment settings used to manage attacks also varied. Following unsatisfactory care experiences at hospitals, some patients reported avoiding further hospital services for later attacks. Therefore, using settings of care as a measure of attack severity should be avoided. Ninety-four percent of patients also experienced chronic symptoms, which were as varied as acute attacks. Pain was the predominant chronic symptom and was managed with opioids in severe cases. Regardless of AAR, porphyria heavily impacted the daily lives of patients and caregivers. Although patients experiencing frequent attacks generally endured a greater impact on their daily life, patients with less frequent attacks also experienced impacts on all domains (social, leisure activities, relationship with family, relationships, psychological wellbeing, finances, employment, and study). Caregivers were most affected in the finance, relationships with family, and employment domains, and just over half of the caregivers reported a moderate impact on their psychological wellbeing., Conclusions/implications: The burden of illness with AHP is high across all patients, regardless of frequency of attacks, and AHP negatively affects patients and caregivers alike.

Successful pregnancy inevitably depends on the implantation of a competent embryo into a receptive endometrium. Although many substances have been suggested to improve the rate of embryo implantation targeting enhancement of endometrial receptivity, currently there rarely are effective evidence-based treatments to prevent or cure this condition. Here we strongly suggest minimally-invasive intra-uterine administration of embryo-secreted chemokine CXCL12 as an effective therapeutic intervention. Chemokine CXCL12 derived from pre- and peri-implanting embryos significantly enhances the rates of embryo attachment and promoted endothelial vessel formation and sprouting in vitro. Consistently, intra-uterine CXCL12 administration in C57BL/6 mice improved endometrial receptivity showing increased integrin β3 and its ligand osteopontin, and induced endometrial angiogenesis displaying increased numbers of vessel formation near the lining of endometrial epithelial layer with higher CD31 and CD34 expression. Furthermore, intra-uterine CXCL12 application dramatically promoted the rates of embryo implantation with no morphologically retarded embryos. Thus, our present study provides a novel evidence that improved uterine endometrial receptivity and enhanced angiogenesis induced by embryo-derived chemokine CXCL12 may aid to develop a minimally-invasive therapeutic strategy for clinical treatment or supplement for the patients with repeated implantation failure with less risk.

There is currently no cure for infertility in women with a poor ovarian response (POR). Neogenin is reported to be abundantly expressed in the ovary; however, its role in mammalian follicular development is unclear and its ligand and signaling pathway remain uncertain. We systematically investigated the role of neogenin and the ligand repulsive guidance molecule c (RGMc) during follicular development. We treated hyperstimulated mouse ovaries with RGMc and analyzed follicular development. Furthermore, we investigated clusters of up/downregulated genes in RGMc-treated ovaries using whole-transcriptome next-generation sequencing (NGS). In addition, we investigated whether expression of up/downregulated factors identified by NGS was also altered in cumulus cells (CCs) of patients with a POR. The number of oocytes was 40% higher in RGMc-treated ovaries than in control ovaries. NGS data indicated that prostaglandin D2 (PGD2) was involved in the RGMc signaling pathway during follicular development. RGMc treatment significantly elevated the PGD2 level in culture medium of CCs obtained from patients with a POR. Our results demonstrate that RGMc as neogenin ligand promotes follicular development in ovaries via the PGD2 signaling pathway. Therefore, it may be possible to use RGMc for ovarian stimulation in patients with a POR.

Background: As the US population ages, safe surgical procedures are necessary for treatment of cutaneous neoplasms in very elderly patients., Objective: To determine the incidence of complications associated with Mohs micrographic surgery (MMS) in patients aged 85 and older, and the risk factors that predispose to complications., Methods: A 9-year retrospective chart review of patients aged 85 and older who underwent MMS at our institution between 07/2007 and 11/2016 was performed. Six types of complications associated with scalpel-based cutaneous surgery were recorded, as well as patient, tumor, and repair characteristics., Results: This study included 949 patients totaling in 1683 MMS cases. There were 30 complications: infection (N = 11), wound dehiscence (N = 6), hematoma (N = 6), hemorrhage (N = 5), flap necrosis (N = 1), and graft necrosis (N = 1), resulting in an overall complication rate of 1.78%. Independent risk factors associated with a statistically higher incidence of complications were anticoagulant use (odds ratio [OR], 2.78; 95% confidence interval [CI], 1.26-6.13; p = .012), extremity location (OR, 2.80; 95% CI, 1.19-6.54; p = .018), greater than 2 MMS stages (OR, 2.43; 95% CI, 1.08-5.46; p = .032), and flap repair (OR, 2.27; 95% CI, 1.05-4.90; p = .036)., Conclusion: Mohs micrographic surgery is a safe procedure for treatment of cutaneous neoplasms in the very elderly., (Copyright © 2020 by the American Society for Dermatologic Surgery, Inc. Published by Wolters Kluwer Health, Inc. All rights reserved.)

Background/Objectives: The serious illness conversation (SIC) is an evidence-based framework for conversations with patients about a serious illness diagnosis. The objective of our study was to develop and validate a novel tool, the SIC-evaluation exercise (SIC-Ex), to facilitate assessment of resident-led conversations with oncology patients. Design: We developed the SIC-Ex based on SIC and on the Royal College of Canada Medical Oncology milestones. Seven resident trainees and 10 evaluators were recruited. Each trainee conducted an SIC with a patient, which was videotaped. The evaluators watched the videos and evaluated each trainee by using the novel SIC-Ex and the reference Calgary-Cambridge guide (CCG) at months zero and three. We used Kane's validity framework to assess validity. Results: Intra-class correlation using average SIC-Ex scores showed a moderate level of inter-evaluator agreement (range 0.523-0.822). Most evaluators rated a particular resident similar to the group average, except for one to two evaluator outliers in each domain. Test-retest reliability showed a moderate level of consistency among SIC-Ex scores at months zero and three. Global rating at zero and three months showed fair to good/very good inter-evaluator correlation. Pearson correlation coefficients comparing total SIC-Ex and CCG scores were high for most evaluators. Self-scores by trainees did not correlate well with scores by evaluators. Conclusions: SIC-Ex is the first assessment tool that provides evidence for incorporating the SIG guide framework for evaluation of resident competence. SIC-Ex is conceptually related to, but more specific than, CCG in evaluating serious illness conversation skills., Competing Interests: No competing financial interests exist., (© Jenny J. Ko et al., 2020; Published by Mary Ann Liebert, Inc.)

Background: Aging has detrimental effects on the ovary, such as a progressive reduction in fertility and decreased hormone production, that greatly reduce the quality of life of women. Thus, the current study was undertaken to investigate whether human placenta-derived mesenchymal stem cell (hPD-MSC) treatment can restore the decreases in folliculogenesis and ovarian function that occur with aging., Methods: Acclimatized 52-week-old female SD rats were randomly divided into four groups: single hPD-MSC (5 × 10 5 ) therapy, multiple (three times, 10-day intervals) hPD-MSC therapy, control (PBS), and non-treated groups. hPD-MSC therapy was conducted by tail vein injection into aged rats. The rats were sacrificed 1, 2, 3, and 5 weeks after the last injection. hPD-MSC tracking and follicle numbers were histologically confirmed. The serum levels of sex hormones and circulating miRNAs were detected by ELISA and qRT-PCR, respectively. TGF-β superfamily proteins and SMAD proteins in the ovary were detected by Western blot analysis., Results: We observed that multiple transplantations of hPD-MSCs more effectively promoted primordial follicle activation and ovarian hormone (E 2 and AMH) production than a single injection. After hPD-MSC therapy, the levels of miR-21-5p, miR-132-3p, and miR-212-3p, miRNAs associated with the ovarian reserve, were increased in the serum. Moreover, miRNAs (miR-16-5p, miR-34a-5p, and miR-191-5p) with known adverse effects on folliculogenesis were markedly suppressed. Importantly, the level of miR-145-5p was reduced after single- or multiple-injection hPD-MSC therapy, and we confirmed that miR-145-5p targets Bmpr2 but not Tgfbr2. Interestingly, downregulation of miR-145-5p led to an increase in BMPR2, and activation of SMAD signaling concurrently increased primordial follicle development and the number of primary and antral follicles., Conclusions: Our study verified that multiple intravenous injections of hPD-MSCs led to improved ovarian function via miR-145-5p and BMP-SMAD signaling and proposed the future therapeutic potential of hPD-MSCs to promote ovarian function in women at advanced age to improve their quality of life during climacterium.

Extracellular particles (EPs) including extracellular vesicles (EVs) and exomeres play significant roles in diseases and therapeutic applications. However, their spatiotemporal dynamics in vivo have remained largely unresolved in detail due to the lack of a suitable method. Therefore, a bioluminescence resonance energy transfer (BRET)-based reporter, PalmGRET, is created to enable pan-EP labeling ranging from exomeres (<50 nm) to small (<200 nm) and medium and large (>200 nm) EVs. PalmGRET emits robust, sustained signals and allows the visualization, tracking, and quantification of the EPs from whole animal to nanoscopic resolutions under different imaging modalities, including bioluminescence, BRET, and fluorescence. Using PalmGRET, it is shown that EPs released by lung metastatic hepatocellular carcinoma (HCC) exhibit lung tropism with varying distributions to other major organs in immunocompetent mice. It is further demonstrated that gene knockdown of lung-tropic membrane proteins, solute carrier organic anion transporter family member 2A1, alanine aminopeptidase/Cd13, and chloride intracellular channel 1 decreases HCC-EP distribution to the lungs and yields distinct biodistribution profiles. It is anticipated that EP-specific imaging, quantitative assays, and detailed in vivo characterization are a starting point for more accurate and comprehensive in vivo models of EP biology and therapeutic design., Competing Interests: The authors declare no conflict of interest., (© 2020 The Authors. Published by Wiley‐VCH GmbH.)

Background: With the growing adoption of the electronic health record (EHR) worldwide over the last decade, new opportunities exist for leveraging EHR data for detection of rare diseases. Rare diseases are often not diagnosed or delayed in diagnosis by clinicians who encounter them infrequently. One such rare disease that may be amenable to EHR-based detection is acute hepatic porphyria (AHP). AHP consists of a family of rare, metabolic diseases characterized by potentially life-threatening acute attacks and chronic debilitating symptoms. The goal of this study was to apply machine learning and knowledge engineering to a large extract of EHR data to determine whether they could be effective in identifying patients not previously tested for AHP who should receive a proper diagnostic workup for AHP., Methods and Findings: We used an extract of the complete EHR data of 200,000 patients from an academic medical center and enriched it with records from an additional 5,571 patients containing any mention of porphyria in the record. After manually reviewing the records of all 47 unique patients with the ICD-10-CM code E80.21 (Acute intermittent [hepatic] porphyria), we identified 30 patients who were positive cases for our machine learning models, with the rest of the patients used as negative cases. We parsed the record into features, which were scored by frequency of appearance and filtered using univariate feature analysis. We manually choose features not directly tied to provider attributes or suspicion of the patient having AHP. We trained on the full dataset, with the best cross-validation performance coming from support vector machine (SVM) algorithm using a radial basis function (RBF) kernel. The trained model was applied back to the full data set and patients were ranked by margin distance. The top 100 ranked negative cases were manually reviewed for symptom complexes similar to AHP, finding four patients where AHP diagnostic testing was likely indicated and 18 patients where AHP diagnostic testing was possibly indicated. From the top 100 ranked cases of patients with mention of porphyria in their record, we identified four patients for whom AHP diagnostic testing was possibly indicated and had not been previously performed. Based solely on the reported prevalence of AHP, we would have expected only 0.002 cases out of the 200 patients manually reviewed., Conclusions: The application of machine learning and knowledge engineering to EHR data may facilitate the diagnosis of rare diseases such as AHP. Further work will recommend clinical investigation to identified patients' clinicians, evaluate more patients, assess additional feature selection and machine learning algorithms, and apply this methodology to other rare diseases. This work provides strong evidence that population-level informatics can be applied to rare diseases, greatly improving our ability to identify undiagnosed patients, and in the future improve the care of these patients and our ability study these diseases. The next step is to learn how best to apply these EHR-based machine learning approaches to benefit individual patients with a clinical study that provides diagnostic testing and clinical follow up for those identified as possibly having undiagnosed AHP., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: GIVLAARI is a product of Alnylam. GIVLAARI is a prescription medicine used to treat acute hepatic porphyria (AHP) in adults. This work was funded and the associated editorial support was provided by Alnylam Pharmaceuticals, Inc., Cambridge, MA. Grant number 4510005336, https://www.alnylam.com/ Alnylam participated in algorithm design and preparation of the manuscript. They had no role in the evaluation or EHR data collection and analysis, nor did they have any access to the individual patient electronic health record data used in this research. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Background: Up-regulation of hepatic delta-aminolevulinic acid synthase 1 (ALAS1), with resultant accumulation of delta-aminolevulinic acid (ALA) and porphobilinogen, is central to the pathogenesis of acute attacks and chronic symptoms in acute hepatic porphyria. Givosiran, an RNA interference therapy, inhibits ALAS1 expression., Methods: In this double-blind, placebo-controlled, phase 3 trial, we randomly assigned symptomatic patients with acute hepatic porphyria to receive either subcutaneous givosiran (2.5 mg per kilogram of body weight) or placebo monthly for 6 months. The primary end point was the annualized rate of composite porphyria attacks among patients with acute intermittent porphyria, the most common subtype of acute hepatic porphyria. (Composite porphyria attacks resulted in hospitalization, an urgent health care visit, or intravenous administration of hemin at home.) Key secondary end points were levels of ALA and porphobilinogen and the annualized attack rate among patients with acute hepatic porphyria, along with hemin use and daily worst pain scores in patients with acute intermittent porphyria., Results: A total of 94 patients underwent randomization (48 in the givosiran group and 46 in the placebo group). Among the 89 patients with acute intermittent porphyria, the mean annualized attack rate was 3.2 in the givosiran group and 12.5 in the placebo group, representing a 74% lower rate in the givosiran group (P<0.001); the results were similar among the 94 patients with acute hepatic porphyria. Among the patients with acute intermittent porphyria, givosiran led to lower levels of urinary ALA and porphobilinogen, fewer days of hemin use, and better daily scores for pain than placebo. Key adverse events that were observed more frequently in the givosiran group were elevations in serum aminotransferase levels, changes in serum creatinine levels and the estimated glomerular filtration rate, and injection-site reactions., Conclusions: Among patients with acute intermittent porphyria, those who received givosiran had a significantly lower rate of porphyria attacks and better results for multiple other disease manifestations than those who received placebo. The increased efficacy was accompanied by a higher frequency of hepatic and renal adverse events. (Funded by Alnylam Pharmaceuticals; ENVISION ClinicalTrials.gov number, NCT03338816.)., (Copyright © 2020 Massachusetts Medical Society.)

Objective: Endometrial fibrosis, the primary pathological feature of intrauterine adhesion, may lead to disruption of endometrial tissue structure, menstrual abnormalities, infertility, and recurrent pregnancy loss. At present, no ideal therapeutic strategy exists for this fibrotic disease. Eupatilin, a major pharmacologically active flavone from Artemisia, has been previously reported to act as a potent inducer of dedifferentiation of fibrotic tissue in the liver and lung. However, the effects of eupatilin on endometrial fibrosis have not yet been investigated. In this study, we present the first report on the impact of eupatilin treatment on transforming growth factor beta (TGF-β)-induced endometrial fibrosis., Methods: The efficacy of eupatilin on TGF-β-induced endometrial fibrosis was assessed by examining changes in morphology and the expression levels of fibrosis markers using immunofluorescence staining and quantitative real-time reverse-transcription polymerase chain reaction., Results: Eupatilin treatment significantly reduced the fibrotic activity of TGF-β-induced endometrial fibrosis in Ishikawa cells, which displayed more circular shapes and formed more colonies. Additionally, the effects of eupatilin on fibrotic markers including alpha-smooth muscle actin, hypoxia-inducible factor 1 alpha, collagen type I alpha 1 chain, and matrix metalloproteinase-2, were evaluated in TGF-β-induced endometrial fibrosis. The expression of these markers was highly upregulated by TGF-β pretreatment and recovered to the levels of control cells in response to eupatilin treatment., Conclusion: Our findings suggest that suppression of TGF-β-induced signaling by eupatilin might be an effective therapeutic strategy for the treatment of endometrial fibrosis.