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The spleen is a compartmentalized organ that serves as a blood filter and safeguard of systemic immune surveillance. Labyrinthine networks of fibroblastic stromal cells construct complex niches within the white pulp and red pulp that are important for tissue homeostasis and immune activation. However, the identity and roles of the global splenic fibroblastic stromal cells in homeostasis and immune responses are poorly defined. Here, we performed a cellular and molecular dissection of the splenic reticular stromal cell landscape. We found that white pulp fibroblastic reticular cells (FRCs) responded robustly during acute viral infection, but this program of gene regulation was suppressed during persistent viral infection. Single-cell transcriptomic analyses in mice revealed diverse fibroblast cell niches and unexpected heterogeneity among podoplanin-expressing cells that include glial, mesothelial, and adventitial cells in addition to FRCs. We found analogous fibroblastic stromal cell diversity in the human spleen. In addition, we identify the transcription factor SpiB as a critical regulator required to support white pulp FRC differentiation, homeostatic chemokine expression, and antiviral T cell responses. Together, our study provides a comprehensive map of fibroblastic stromal cell types in the spleen and defines roles for red and white pulp fibroblasts for splenic function and orchestration of immune responses.

Increased viral risk donors (IVRDs) with increased risk behaviors for blood-borne virus infection and negative nucleic acid testing have a low absolute risk of "window period" infection. Utilization and allocation of IVRD organs differ between jurisdictions., Methods: We examined the characteristics and utilization of deceased donor IVRD kidneys and recipient outcomes within a 2-y period (July 31, 2018-July 31, 2020) postimplementation of a new opt-in allocation pathway for preconsented recipients in Victoria, Australia., Results: Fifty-six kidneys from 31 IVRDs were utilized, comprising 13% of donors. Preconsent rate to accept IVRD kidneys increased to 41% of the waitlist in the 2 y postimplementation, and IVRDs having no kidneys utilized reduced to 0%. Compared with non-IVRD kidneys, kidney offer declines >10 per donor were less likely from IVRDs (3% vs 19%; P < 0.05). IVRDs were younger (median age 36 [IQR 30-44] vs 51 [35-60] y; P < 0.0001), with lower kidney donor profile index (25% [13-40%] vs 57% [29-75%]; P < 0.0001), and less hypertension (0% vs 22%; P < 0.01). Estimated glomerular filtration rate 3 mo post-transplant was superior ( P < 0.01). Injecting drug use (61%) was the most common increased risk behavior. 29% of IVRDs were hepatitis C antibody positive but nucleic acid testing negative. No active infection was detected in any recipient post-transplant., Conclusions: The described opt-in system permits efficient allocation and utilization of kidneys from IVRDs, with superior quality and graft function. Education is crucial to facilitate informed consent and equity of access to this donor pool., Competing Interests: The authors declare no funding or conflicts of interest., (Copyright © 2021 The Author(s). Transplantation Direct. Published by Wolters Kluwer Health, Inc.)

Background: Even in the extended-criteria era, the reasons for declining lung donors are not always clear. Furthermore, it has not been determined how many actual declined lungs would be retrieved by ex vivo lung perfusion (EVLP) beyond that already achieved in centers with an existing high utilization rate., Methods: This retrospective study reviewed all lung donor referrals between 2014 and 2018, including detailed formal referrals and preliminary notifications. This study categorized reasons for lung donor non-acceptance and estimated how many declined grafts could have been theoretically retrievable by using EVLP., Results: In total, 966 lung donor candidates were referred, including 313 transplanted donors, 336 declined donors after detailed referrals (group A) and 258 preliminary declined. In group A, the primary reasons for refusal were lung quality issues (49%), general medical issues (25%), and organization issues (26%), combined with secondary reasons in many cases. Main lung quality issues were an extensive smoking history, abnormal chest radiography, and underlying lung disease. Although 73 declined lung donors had indications for EVLP, the retrievable lungs decreased to only 30 cases after considering the details of all clinical contraindications and organizational issues. Nevertheless, 59 intended donation after circulatory death donors did not progress to death after withdrawal of cardiorespiratory support in the required timeframe, and EVLP may have an emerging additional role here., Conclusions: Based on commonly cited criteria for EVLP indication, the number of EVLP retrievable lung donors represented only a small portion of declined donor lungs referred to our center from the state donation network., (Copyright © 2021 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

Although the use of donation after circulatory death (DCD) donors has increased lung transplant activity, 25-40% of intended DCD donors do not convert to actual donation because of no progression to asystole in the required time frame after withdrawal of cardiorespiratory support (WCRS). No studies have specifically focussed on DCD lung donor progression. This retrospective study reviewed intended DCD lung donors to make a prediction model of the likelihood of progression to death using logistic regression and classification and regression tree (CART). Between 2014 and 2018, 159 of 334 referred DCD donors were accepted, with 100 progressing to transplant, while 59 (37%) did not progress. In logistic regression, a length of ICU stay ≤ 5 days, severe infra-tentorial brain damage on imaging and use of vasopressin were related with the progression to actual donation. CART modelling of the likelihood of death within 90-minute post-WCRS provided prediction with a sensitivity of 1.00 and positive predictive value of 0.56 in the validation data set. In the nonprogressed DCD group, 26 died within 6 h post-WCRS. Referral received early after ICU admission, with nonspontaneous ventilatory mode, deep coma and severe infra-tentorial damage were relevant predictors. The CART model is useful to exclude DCD donor candidates with low probability of progression., (© 2021 Steunstichting ESOT. Published by John Wiley & Sons Ltd.)

Background: Although it is intense in health care resources, by facilitating assessment and reconditioning, ex vivo lung perfusion (EVLP) has the potential to expand the donor pool and improve lung transplant outcomes. However, inclusion criteria used in EVLP trials have not been validated., Methods: This retrospective study from 2014 to 2018 reviewed our local state-based donation organization donor records as well as subsequent recipient outcomes to explore the relation between EVLP indications used in clinical trials and recipient outcomes. The primary outcome was primary graft dysfunction grade 3 at 24 hours, with 30-day mortality and posttransplant survival time as secondary outcomes, compared with univariate and multivariate analysis., Results: From 705 lung donor referrals, 304 lung transplantations were performed (use rate of 42%); 212 of recipients (70%) met at least 1 of the commonly cited EVLP initiation criteria. There was no significant difference in primary graft dysfunction grade 3 or 30-day mortality between recipients with or without an EVLP indication (10.2% versus 7.8%, P = .51; and 2.4% versus 0%, P = .14, respectively). Multivariate analyses showed no significant relationship between commonly cited EVLP criteria and primary graft dysfunction grade 3 or survival time. Recipient outcomes were significantly associated with recipient diagnosis., Conclusions: At least 1 commonly cited criterion for EVLP initiation was present in 70% of the transplanted donors, and yet it did not predict clinical results; acceptable outcomes were seen in both subgroups. To discover the true utility of EVLP beyond good clinical management and focus EVLP on otherwise unacceptable lungs, a reconsideration of EVLP inclusion criteria is required., (Copyright © 2021 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

Objectives: Endemic human coronaviruses (hCoVs) circulate worldwide but cause minimal mortality. Although seroconversion to hCoV is near ubiquitous during childhood, little is known about hCoV-specific T-cell memory in adults., Methods: We quantified CD4 T-cell and antibody responses to hCoV spike antigens in 42 SARS-CoV-2-uninfected individuals. Antigen-specific memory T cells and circulating T follicular helper (cTFH) cells were identified using an activation-induced marker assay and characterised for memory phenotype and chemokine receptor expression., Results: T-cell responses were widespread within conventional memory and cTFH compartments but did not correlate with IgG titres. SARS-CoV-2 cross-reactive T cells were observed in 48% of participants and correlated with HKU1 memory. hCoV-specific T cells exhibited a CCR6 + central memory phenotype in the blood, but were enriched for frequency and CXCR3 expression in human lung-draining lymph nodes., Conclusion: Overall, hCoV-specific humoral and cellular memory are independently maintained, with a shared phenotype existing among coronavirus-specific CD4 T cells. This understanding of endemic coronavirus immunity provides insight into the homeostatic maintenance of immune responses that are likely to be critical components of protection against SARS-CoV-2., Competing Interests: The authors declare no conflict of interest., (© 2021 The Authors. Clinical & Translational Immunology published by John Wiley & Sons Australia, Ltd on behalf of Australian and New Zealand Society for Immunology Inc.)

Background: Donor smoking histories are common in the lung donor pool, which are known to adversely affect post-lung transplant (LTx) outcomes. However, no evidence is available about smoking status (current/former), cumulative dose effect, or the combined effect of tobacco with marijuana., Methods: We retrospectively reviewed our local state-based donation organization records and subsequent LTx recipient outcomes. The primary outcome was 3-year graft survival, with cause of death as secondary outcomes. Univariate and multivariate Cox regression analyses were used to explore smoking status or cumulative dose effect., Results: Between 2014 and 2018, 304 LTxs were performed: 133 (44%) LTxs were from never-smoker donors, 68 (22%) from former-smoker donors, and 103 (34%) from current-smoker donors. Of the current-smoker donors, 48% had a marijuana use history. There was no significant difference in early mortality, although recipients who received transplants from current-smoker donors had a lower 3-year graft survival than those who received transplants from never smokers. Multivariate modeling showed that current tobacco smoking (hazard ratio: 2.13, 95% CI: 1.13-3.99) and a more than 5-year weekly marijuana use (hazard ratio: 2.97, 95% CI: 1.29-6.87) were independent donor factors affecting graft survival. Chronic lung allograft dysfunction accounted for a higher proportion of the causes of death within 3 years after LTx where lungs from current/former smokers were utilized compared with those from never smokers (chronic lung allograft dysfunction-cause mortality: 11%, 7%, 0%, respectively)., Conclusions: More than 50% of LTx donors had smoking histories. Current tobacco use or more than 5-year weekly marijuana smoking history adversely affected 3-year graft survival. Our findings support the importance of obtaining a detailed donor tobacco and marijuana smoking history., (Copyright © 2020 International Society for Heart and Lung Transplantation. Published by Elsevier Inc. All rights reserved.)

Purpose: Prophylactic laxative regimens may prevent constipation but may increase diarrhea and subsequent rectal tube insertion. Our aim was to compare three prophylactic laxative regimens on the rate of rectal tube insertion (primary outcome) and major constipation- or diarrhea-associated complications., Material and Methods: We conducted a cluster-crossover trial. Three pods in a single ICU were each randomized to one of three regimens for four months with rolling cross-over. All mechanically-ventilated and enterally-fed adult patients received either regimen: A) one coloxyl with senna BD from day one; B) two coloxyl with senna +20 ml lactulose BD commencing on day 3; or C) two coloxyl with senna tablets +20 ml lactulose BD commencing on day 6., Results: We enrolled 570 patients (A = 170, B = 205, C = 195) with similar baseline features. Overall, 53 (9.3%) patients received a rectal tube, and insertion rate was not statistically different between the three regimens (A = 12.9%, B = 7.8%, C = 7.7%; p = 0.15). The proportions of patients with other major constipation- or diarrhea-associated complications were similar, as were major patient-centred outcomes., Conclusion: Earlier commencement of a prophylactic coloxyl-based laxative regimen (day 1 or 3) did not affect the rates of complications associated with constipation or diarrhea when compared to delayed introduction (day 6)., (Copyright © 2019. Published by Elsevier Inc.)

The aim of this multicenter, prospective, observer-blinded, parallel group, randomized controlled trial was to assess the safety and efficacy of EDX110, a nitric oxide generating medical device, in the treatment of diabetic foot ulcers in a patient group reflecting "real world" clinical practice compared against optimal standard care. Participants were recruited from ten hospital sites in multidisciplinary foot ulcer clinics. The ulcers were full thickness, with an area of 25-2,500 mm 2 and either a palpable pedal pulse or ankle brachial pressure index > 0.5. Infected ulcers were included. Treatment lasted 12 weeks, or until healed, with a 12-week follow-up period. Both arms were given optimal debridement, offloading and antimicrobial treatment, the only difference being the fixed used of EDX110 as the wound dressing in the EDX110 group. 135 participants were recruited with 148 ulcers (EDX110-75; Control-73), 30% of which were clinically infected at baseline. EDX110 achieved its primary endpoint by attaining a median Percentage Area Reduction of 88.6% compared to 46.9% for the control group (p = 0.016) at 12 weeks in the intention-to-treat population. There was no significant difference between wound size reduction achieved by EDX110 after 4 weeks and the wound size reduction achieved in the control group after 12 weeks. EDX110 was well tolerated. Thirty serious adverse events were reported (12 in the EDX110 group, of which 4 were related to the ulcer; 18 in the control group, of which 10 were related and 1 possibly related to the ulcer), with significant reduction in serious adverse events related to the ulcer in EDX group. There was no significant difference in adverse events. This study, in a real world clinical foot ulcer population, demonstrates the ability of EDX110 to improve healing, as measured by significantly reducing the ulcer area, compared to current best clinical practice., (© 2018 by the Wound Healing Society.)

Aim: Organ and tissue donation (OTD) rates in New Zealand are low compared to many countries. Young adults are 'tomorrow's donors', yet the attitudes and knowledge of this group to OTD have not been examined locally. Such information is relevant to ODT education and clinical engagement., Method: A random sample of University of Otago students (<25 years, permanent New Zealand resident) was surveyed to examine OTD knowledge and attitudes. This included general knowledge, OTD policy (opt-in, opt-out), donation by self, and donation by loved ones. Questions included yes-no, multiple choice, and Likert-type responses. Analyses by sex, demographic characteristics, supportive attitudes to ODT, and University of Otago student profile were performed., Results: 180 responses were gathered (mean age 20.1 years, 67% female, 68% New Zealand European); there were no age or response differences between sexes, participants were generally not representative of the University of Otago student profile. Outcomes indicated limited OTD knowledge, positive support for OTD, and willingness to engage in donation the decision-making process for loved ones. Differences between supportive and non-supportive OTD attitudes was seen for some questions., Conclusion: Findings highlight areas for strategic OTD public engagement and provide details relevant to guiding appropriate clinical interaction in facilitating decisions about OTD.

Background: Alpha-methylacyl-coA racemase (AMACR) was recently shown to be a sensitive immunohistochemical marker for substantiating a diagnosis of adenocarcinoma of the prostate. Its applicability to exfoliative urinary cytology has not been investigated before., Case: The patient was a 77-year-old male with a history of persistent intermittent hematuria who was evaluated with urine cytology. Cytopathologic studies were interpreted to demonstrate "atypical urothelial cells." The patient was evaluated with computed tomography, which demonstrated new left hydroureter and hydronephrosis. He underwent cystoscopy, which showed an abnormal left hemitrigone with the left ureteral orifice obscured by an infiltrative mass. This area was biopsied, and histologic studies of the tissue chips demonstrated underlying prostatic adenocarcinoma directly invading the urothelium. We used AMACR immunoreactivity on a retrospectively studied, catheterized urine slide to confirm the diagnosis., Conclusion: This case suggests that combined cytomorphology and immunocytochemisty for AMACR may allow an accurate identification of cells of prostatic adenocarcinoma when cytomorphologic studies or the clinical history raises the differential diagnosis of prostate cancer presenting with exfoliation of malignant cells into the urine.

Crystallizations of pharmaceutical active ingredients, particularly those that posses multiple polymorphic forms, are among the most critical and least understood pharmaceutical manufacturing processes. Many process and product failures can be traced to a poor understanding and control of crystallization processes. The Food and Drug Administration's process analytical technology (PAT) initiative is a collaborative effort with industry to introduce new and efficient manufacturing technologies into the pharmaceutical industry. PAT's are systems for design, analysis, and control of manufacturing processes. They aim to assure high quality through timely measurements of critical quality and performance attributes of raw materials, in-process materials, and final products. Implementation of PAT involves scientifically based process design and optimization, appropriate sensor technologies, statistical and information tools (chemometrics), and feedback process control strategies working together to produce quality products. This review introduces the concept of PAT and discusses its application to crystallization processes through review of several case studies. A variety of in situ analytical methods combined with chemometric tools for analysis of multivariate process information provide a basis for future improvements in modeling, simulation, and control of crystallization processes.

Detailed invasive haemodynamic studies were performed in 27 of 32 patients with severe tetanus. Nineteen had severe uncomplicated tetanus and eight had associated major complications, chiefly infection and pulmonary complications. The results were compared with those obtained from 15 healthy male volunteers who served as controls. There were two deaths in 32 patients (mortality 6.25 per cent). Severe tetanus without major complications was characterized by a high output hyperkinetic circulatory state with tachycardia (heart rate 131 (19.2) beats/minute), increased stroke volume index (43.1 (10.7) ml/m2), increased cardiac index (5.48 (0.94) l/min/m2) and a normal left ventricular stroke work index (60.5 (15.9) g/m/m2). Volume loading demonstrated a significant haemodynamic response and increased vascular capacitance. Even so the maximum percent rise from baseline values of these indices after volume load was significantly higher in controls (p < 0.001). Autonomic cardiovascular disturbances affected both sympathetic and parasympathetic activity. Hypertension and tachycardia alternating with hypotension and bradycardia were related to sudden fluctuations in systemic vascular resistance. Our studies suggested some degree of myocardial dysfunction in patients with severe uncomplicated tetanus. The haemodynamics of severe tetanus were masked and altered by complicating infection, pneumonia, and atelectasis.