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Abstract Objectives In response to COVID-19’s first wave, provincial governments rapidly implemented several public health directives, including isolation measures and care facility visitor restrictions, which profoundly affected healthcare delivery at the end of life and dying experiences and perceptions. The objective of this study was to identify implications of early policy changes for dying at home. Methods Analysis of interviews with 29 key informants with expertise in the policy and practice context of dying at home and care for those dying at home was conducted as part of a larger mixed-methods study on dying at home in Canada. Results Initial pandemic policy responses, especially visitor restrictions and limitations to home care services, shaped dying at home in relation to three themes: (1) increasing preferences and demand for, yet constrained system ability to support dying at home; (2) reinforcing and illuminating systemic reliance on and need for family/friend caregivers and community organizations, while constraining their abilities to help people die at home; and (3) illuminating challenges in developing and implementing policy changes during a pandemic, including equity-related implications. Conclusion This study contributes to broader understanding of the multifaceted impacts of COVID-19 policy responses in various areas within Canadian healthcare systems. Implications for healthcare delivery and policy development include (1) recognizing the role of family/friend caregivers and community organizations in end-of-life care, (2) recognizing health inequities at the end of life, and (3) considering possible changes in future end-of-life preferences and public attitudes about dying at home and responsibility for end-of-life care.

A leading cause of mortality after influenza infection is the development of a secondary bacterial pneumonia. In the absence of a bacterial superinfection, prescribing antibacterial therapies is not indicated but has become a common clinical practice for those presenting with a respiratory viral illness. In a murine model, we found that antibiotic use during influenza infection impaired the lung innate immunologic defenses toward a secondary challenge with methicillin-resistant Staphylococcus aureus (MRSA). Antibiotics augment lung eosinophils, which have inhibitory effects on macrophage function through the release of major basic protein. Moreover, we demonstrated that antibiotic treatment during influenza infection caused a fungal dysbiosis that drove lung eosinophilia and impaired MRSA clearance. Finally, we evaluated 3 cohorts of hospitalized patients and found that eosinophils positively correlated with antibiotic use, systemic inflammation, and worsened outcomes. Altogether, our work demonstrates a detrimental effect of antibiotic treatment during influenza infection that has harmful immunologic consequences via recruitment of eosinophils to the lungs, thereby increasing the risk of developing a secondary bacterial infection.

Single-cell RNA sequencing (scRNA-seq) of peripheral blood mononuclear cells (PBMCs) allows in-depth assessment of transcriptional changes in immune cells of patients with COVID-19. However, collecting, processing, and analyzing samples from patients with COVID-19 pose many challenges because blood samples may contain infectious virus, identification of immune cell subtypes can be difficult, and biological interpretation of analytical results is complex. To address these issues, we describe a protocol for sample processing, sorting, methanol fixation, and scRNA-seq analysis of PBMCs from frozen buffy coat samples from patients with COVID-19. For complete details on the use and execution of this protocol, please refer to (Yao et al., 2021)., Competing Interests: The authors declare no competing interests., (© 2021 The Authors.)

Idiopathic pulmonary fibrosis (IPF) is a chronic and fatal lung disease. A maladaptive epithelium due to chronic injury is a prominent feature and contributor to pathogenic cellular communication in IPF. Recent data highlight the concept of a "reprogrammed" lung epithelium as critical in the development of lung fibrosis. Extracellular vesicles (EVs) are potent mediator of cellular crosstalk, and recent evidence supports their role in lung pathologies such as IPF. Here, we demonstrate that syndecan-1 is overexpressed by the epithelium in the lungs of IPF patients and in murine models after bleomycin injury. Moreover, we find that syndecan-1 is a pro-fibrotic signal that alters alveolar type II (ATII) cell phenotypes by augmenting TGFβ and Wnt signaling among other pro-fibrotic pathways. Importantly, we demonstrate that syndecan-1 controls the packaging of several anti-fibrotic microRNAs into EVs that have broad effects over several fibrogenic signaling networks as a mechanism of regulating epithelial plasticity and pulmonary fibrosis. Collectively, our work reveals new insight into how EVs orchestrate cellular signals that promote lung fibrosis and demonstrate the importance of syndecan-1 in coordinating these programs.

Citrobacter rodentium is a gastrointestinal infection that requires early IL-22 from group 3 innate lymphoid cells (ILC3) for resistance. The role of vitamin D in the clearance of C. rodentium infection was tested in vitamin D sufficient (D+) and vitamin D deficient (D-) wildtype (WT) and Cyp27B1 (Cyp) KO mice (unable to produce the high affinity vitamin D ligand 1,25(OH) 2 D, 1,25D). Feeding Cyp KO mice D- diets reduced vitamin D levels and prevented synthesis of 1,25D. D- (WT and Cyp KO) mice had fewer ILC3 cells and less IL-22 than D+ mice. D- Cyp KO mice developed a severe infection that resulted in the lethality of the mice by d14 post-infection. T and B cell deficient D- Rag KO mice also developed a severe and lethal infection with C. rodentium compared to D+ Rag KO mice. D- WT mice survived the infection but took significantly longer to clear the C. rodentium infection than D+ WT or D+ Cyp KO mice. Treating infected D- Cyp KO mice with IL-22 protected the mice from lethality. Treating the D- WT mice with 1,25D reconstituted the ILC3 cells in the colon and protected the mice from C. rodentium . IL-22 treatment of D- WT mice eliminated the need for vitamin D to clear the C. rodentium infection. Vitamin D is required for early IL-22 production from ILC3 cells and protection from enteric infection with C. rodentium .

Vitamin D, 25hydroxyvitamin D (25D), and 24,25dihydroxyvitamin D (24,25D) were measured before and after broad spectrum antibiotic (Abx) treatment for 2 wks. Abx treatments increased 25D and 24,25D levels suggesting that the microbiota or Abx were altering vitamin D metabolism. Increased 25D, but not 24,25D, following Abx treatments were found to be dependent on toll like receptor signaling. Conversely, the effects of Abx on 24,25D levels required that the vitamin D receptor (VDR) be expressed in tissues outside of the hematopoietic system (kidney) and not the immune system. Fibroblast growth factor (FGF)23 increased following Abx treatment and the effect of Abx treatment on FGF23 (like the effect on 24,25D) was not present in VDR knockout (KO) mice. The Abx mediated increase in 24,25D was due to changes to the endocrine regulation of vitamin D metabolism. Conversely, 25D levels went up with Abx treatment of the VDR KO mice. Host sensing of microbial signals regulates the levels of 25D in the host., (Copyright © 2018. Published by Elsevier Inc.)

To determine the effect of the microbiota on vitamin D metabolism, serum 25-hydroxyvitamin D(25D), 24,25-dihydroxyvitamin D (24,25D), and 1,25-dihydroxyvitamin D (1,25D) were measured in germ-free (GF) mice before and after conventionalization (CN). GF mice had low levels of 25D, 24,25D, and 1,25D and were hypocalcemic. CN of the GF mice with microbiota, for 2 weeks recovered 25D, 24,25D, and 1,25D levels. Females had more 25D and 24,25D than males both as GF mice and after CN. Introducing a limited number of commensals (eight commensals) increased 25D and 24,25D to the same extent as CN. Monocolonization with the enteric pathogen Citrobacter rodentium increased 25D and 24,25D, but the values only increased after 4 weeks of C. rodentium colonization when inflammation resolved. Fibroblast growth factor (FGF) 23 was extremely high in GF mice. CN resulted in an increase in TNF-α expression in the colon 2 days after CN that coincided with a reduction in FGF23 by 3 days that eventually normalized 25D, 24,25D, 1,25D at 1-week post-CN and reinstated calcium homeostasis. Neutralization of FGF23 in GF mice raised 1,25D, without CN, demonstrating that the high FGF23 levels were responsible for the low calcium and 1,25D in GF mice. The microbiota induce inflammation in the GF mice that inhibits FGF23 to eventually reinstate homeostasis that includes increased 25D, 24,25D, and 1,25D levels. The microbiota through FGF23 regulates vitamin D metabolism.

Aims: To investigate whether a vaginal speculum examination (VSE) prior to a transvaginal scan (TVS) alters the diagnosis or management of women who complain of bleeding in early pregnancy., Methods: A prospective study. Women were asked to describe their bleeding as heavy, moderate or light and to consent to VSE prior to TVS. TVS was performed to obtain a final diagnosis of pregnancy outcome., Results: 221 consecutive women were included in the study. In 14.5% (n = 32) complaining of heavy bleeding, blood was seen in all but two VSE and 84% (n = 27) had a miscarriage diagnosed by TVS. Products of conception were removed in 18.8% (n = 6), but this did not alter the subsequent immediate management of any cases. 65.2% (n = 144) of women complained of light bleeding, blood was seen on VSE in 53% (n = 77). Of these women, 25% (n = 19) of those where blood was seen had a miscarriage, compared to 6% (n = 4) of women where blood was not seen. A cervical ectropion was visualised in 11.7% (n = 26) and 2.3% (n = 5) had a cervical polyp. No other clinically significant pathology was detected., Conclusion: The amount of bleeding reported by women in early pregnancy relates well with VSE findings. Performing a VSE did not alter the subsequent management of these patients. This study demonstrates that routine objective assessment of blood by a clinician performing VSE prior to a TVS is unnecessary.

Human chorionic gonadotrophin (hCG) is released within hours of fertilization and has a profound ability to downregulate maternal cellular immunity against trophoblastic paternal antigens. It also promotes angiogenic activity of the extravillous trophoblast, and impairment of this function may lead to inadequate placentation and an increased risk of preeclampsia. There is increasing evidence that hCG alters the activity of dendritic cells via an upregulation of indoleamine 2,3-dioxygenase activity. This reduces T-cell activation and cytokine production, as well as encouraging Treg cell recruitment to the fetal-maternal interface. These changes are critical in promoting maternal tolerance. hCG is also able to increase the proliferation of uterine natural killer cells, while reducing the activity of cytotoxic peripheral blood natural killer cells. There are rare reports of autoantibodies directed against hCG or the luteinizing hormone/hCG receptor in women with recurrent miscarriage. These autoantibodies are more frequent in women with thyroid autoimmunity. This may explain the association between thyroid autoimmunity and impaired fertility. Downregulating these anti-hCG and anti-luteinizing hormone/hCG receptor autoantibodies may be helpful in some women with early miscarriage or recurrent failed in vitro fertilization.

The authors report a rare, unusual case of an isolated fallopian tube torsion in a young woman presenting with acute lower abdominal pain and a negative pregnancy test. The patient who initially presented with clinical and biochemical features suggestive of acute appendicitis was found to have a large necrotic pelvic mass at laparoscopy, discovered to be a torted fallopian tube with no ipsilateral ovarian involvement. The patient had a laparoscopic removal of the necrotic tube and recovered well postoperatively.

Background: Persistent trophoblast is a recognised complication of salpingostomy for the treatment of ectopic pregnancy, with reported rates of 3-20%; hence, women are advised to have serum human chorionic gonadotrophin (hCG) levels monitored post-operatively. Although much less common, there are also reports of disseminated trophoblastic peritoneal implants after laparoscopic salpingectomy. The aim of this study was to assess whether monitoring of post-operative serum hCG levels is necessary in women undergoing salpingectomy, where intra-operative spillage of trophoblast is thought to have occurred., Methods: This was a retrospective study of women who underwent serum hCG follow-up after salpingectomy. Serum hCG levels were monitored if: (1) the ectopic pregnancy was found to be ruptured; (2) there was a significant haemoperitoneum (>500 ml); (3) there was thought to be spillage of trophoblast at the time of salpingectomy or (4) a tubal miscarriage was diagnosed. Serum hCG levels were taken at days 1-2, days 3-4, days 6-8 or days 13-15 post-surgery. Women were followed up until the serum hCG level was <15 IU/l. Persistent trophoblast was defined as a failure of the serum hCG level to decrease spontaneously after surgery., Results: 105 women underwent serum hCG follow-up after a laparoscopy for a tubal ectopic pregnancy. Of these women, 92 had a laparoscopic salpingectomy and 13 were diagnosed with a tubal miscarriage at the time of laparoscopy. In all women the serum hCG decreased spontaneously., Conclusion: It does not appear necessary to routinely monitor serum hCG levels post-operatively in women diagnosed with tubal miscarriages, in those undergoing salpingectomy for a ruptured ectopic pregnancy or in cases of salpingectomy, where there is thought to be spillage of trophoblast., (Copyright © 2010 S. Karger AG, Basel.)

Objectives: To assess whether early fetal growth restriction in a twin, expressed as the intertwin discrepancy in crown-rump length (CRL) between two viable twins at 7 + 0 to 9 + 6 weeks' gestation, is predictive of subsequent single fetal loss., Methods: This was a retrospective analysis of data collected prospectively over 3 years. Women attending an early pregnancy unit underwent a transvaginal ultrasound examination. In women with a viable twin pregnancy between 7 + 0 and 9 + 6 weeks of gestation, the CRL ratio between the two embryos was calculated. Women were followed up and the intertwin discrepancy in CRL at the 7 + 0 to 9 + 6-week scan was compared between cases in which there was spontaneous reduction to a singleton and those in which both twins remained viable at the 11-14-week scan., Results: There were 77 women included in the study and nine (12%) of these had a single fetal loss. Pregnancies with subsequent single fetal loss were more likely to have a larger median CRL discrepancy (42.0%; interquartile range (IQR), 23.8-64.3%) than were those which retained two viable fetuses (6.1%; IQR, 2.2-12.5%) (P < 0.0001). The median CRL discrepancy in subsequently viable monochorionic diamniotic twins (10.9%; IQR, 1.9-17.5%) was no different from that in dichorionic diamniotic twin pregnancies (5.9%; IQR, 2.1-12.3%) (P = 0.305). Regardless of chorionicity, there was a relationship between increasing CRL discrepancy and single fetal loss; the likelihood of a subsequently viable twin pregnancy was 97% if the discrepancy was < 20%, while if the discrepancy was > 60% there were no cases of both twins remaining viable (P < 0.0001). Receiver-operating characteristics curve analysis of CRL discrepancy in predicting single fetal loss gave an area under the curve of 0.93, with an optimum cut-off point of 16.9% discrepancy (sensitivity, 88.9%; 95% CI, 51.8-99.7; and specificity, 86.7%; 95% CI, 76.3-93.8)., Conclusion: There is a significant relationship between CRL discrepancy at 7 + 0 to 9 + 6 weeks and the likelihood of subsequent single intrauterine fetal loss. This suggests that spontaneous fetal demise of one twin may be preceded by growth restriction in the first trimester., ((c) 2009 ISUOG.)

Objectives: To assess the agreement in the diagnosis of chorionicity and amnionicity between transvaginal ultrasound examination at 7-9 weeks' gestation and at the 11-14-week transabdominal scan., Methods: This was a retrospective analysis of prospectively collected data of twin pregnancies over a 3-year period. Chorionicity and amnionicity were assessed in women with viable twin pregnancies who underwent a transvaginal ultrasound scan at between 7 and 9 weeks' gestation. These findings were compared with the diagnoses of chorionicity and amnionicity at the subsequent 11-14-week scan in these women., Results: Chorionicity and amnionicity were documented in 67 viable twin pregnancies at both 7-9 and 11-14 weeks' gestation. There was agreement in the chorionicity and amnionicity reported at each of the two scans in 65 out of 67 (97%) cases. Of the dichorionic-diamniotic (DCDA) pregnancies reported at 7-9 weeks, 53 out of 54 (98%) were confirmed at the 11-14-week scan and one (2%) was found to be monochorionic-diamniotic (MCDA). However, at birth these twins were of different sex, confirming DCDA twins as initially diagnosed at 7-9 weeks. Of the 12 pregnancies diagnosed as MCDA at 7-9 weeks, all were found to be MCDA at the 11-14-week scan. There was one monochorionic-monoamniotic (MCMA) pregnancy diagnosed at 7-9 weeks that was subsequently found to be MCDA at the 11-14-week scan., Conclusion: Transvaginal ultrasound examination at 7-9 weeks' gestation shows very high agreement with the 11-14-week scan in the diagnosis of chorionicity and amnionicity in twin pregnancies, suggesting that it provides a similar level of accuracy. Accuracy may be higher for DC twins than MC twins, which may relate to the gestational age at which the sonographic appearance of the amniotic sac develops.