Structure and Function of SNM1 Family Nucleases.

Three human nucleases, SNM1A, SNM1B/Apollo, and SNM1C/Artemis, belong to the SNM1 gene family. These nucleases are involved in various cellular functions, including homologous recombination, nonhomologous end-joining, cell cycle regulation, and telomere maintenance. These three proteins share a similar catalytic domain, which is characterized as a fused metallo-ß-lactamase and a CPSF-Artemis-SNM1-PSO2 domain. SNM1A and SNM1B/Apollo are exonucleases, whereas SNM1C/Artemis is an endonuclease. This review contains a summary of recent research on SNM1's cellular and biochemical functions, as well as structural biology studies. In addition, protein structure prediction by the artificial intelligence program AlphaFold provides a different view of the proteins' non-catalytic domain features, which may be used in combination with current results from X-ray crystallography and cryo-EM to understand their mechanism more clearly.

Red Blood Cell Transfusion in the Intensive Care Unit.

Importance: Red blood cell (RBC) transfusion is common among patients admitted to the intensive care unit (ICU). Despite multiple randomized clinical trials of hemoglobin (Hb) thresholds for transfusion, little is known about how these thresholds are incorporated into current practice. Objective: To evaluate and describe ICU RBC transfusion practices worldwide. Design, Setting, and Participants: International, prospective, cohort study that involved 3643 adult patients from 233 ICUs in 30 countries on 6 continents from March 2019 to October 2022 with data collection in prespecified weeks. Exposure: ICU stay. Main Outcomes and Measures: The primary outcome was the occurrence of RBC transfusion during ICU stay. Additional outcomes included the indication(s) for RBC transfusion (consisting of clinical reasons and physiological triggers), the stated Hb threshold and actual measured Hb values before and after an RBC transfusion, and the number of units transfused. Results: Among 3908 potentially eligible patients, 3643 were included across 233 ICUs (median of 11 patients per ICU [IQR, 5-20]) in 30 countries on 6 continents. Among the participants, the mean (SD) age was 61 (16) years, 62% were male (2267/3643), and the median Sequential Organ Failure Assessment score was 3.2 (IQR, 1.5-6.0). A total of 894 patients (25%) received 1 or more RBC transfusions during their ICU stay, with a median total of 2 units per patient (IQR, 1-4). The proportion of patients who received a transfusion ranged from 0% to 100% across centers, from 0% to 80% across countries, and from 19% to 45% across continents. Among the patients who received a transfusion, a total of 1727 RBC transfusions were administered, wherein the most common clinical indications were low Hb value (n = 1412 [81.8%]; mean [SD] lowest Hb before transfusion, 7.4 [1.2] g/dL), active bleeding (n = 479; 27.7%), and hemodynamic instability (n = 406 [23.5%]). Among the events with a stated physiological trigger, the most frequently stated triggers were hypotension (n = 728 [42.2%]), tachycardia (n = 474 [27.4%]), and increased lactate levels (n = 308 [17.8%]). The median lowest Hb level on days with an RBC transfusion ranged from 5.2 g/dL to 13.1 g/dL across centers, from 5.3 g/dL to 9.1 g/dL across countries, and from 7.2 g/dL to 8.7 g/dL across continents. Approximately 84% of ICUs administered transfusions to patients at a median Hb level greater than 7 g/dL. Conclusions and Relevance: RBC transfusion was common in patients admitted to ICUs worldwide between 2019 and 2022, with high variability across centers in transfusion practices.

Optimization of ether and aniline based inhibitors of lactate dehydrogenase.

Lactate dehydrogenase (LDH) is a critical enzyme in the glycolytic metabolism pathway that is used by many tumor cells. Inhibitors of LDH may be expected to inhibit the metabolic processes in cancer cells and thus selectively delay or inhibit growth in transformed versus normal cells. We have previously disclosed a pyrazole-based series of potent LDH inhibitors with long residence times on the enzyme. Here, we report the elaboration of a new subseries of LDH inhibitors based on those leads. These new compounds potently inhibit both LDHA and LDHB enzymes, and inhibit lactate production in cancer cell lines.

Preoperative identification of cardiac surgery patients at risk of receiving a platelet transfusion: The Australian Cardiac Surgery Platelet Transfusion (ACSePT) risk prediction tool.

Platelet (PLT) transfusions are limited and costly resources. Accurately predicting clinical demand while limiting product wastage remains difficult. A PLT transfusion prediction score was developed for use in cardiac surgery patients who commonly require PLT transfusions. STUDY DESIGN AND METHODS: Using the Australian and New Zealand Society of Cardiac and Thoracic Surgeons National Cardiac Surgery Database, significant predictors for PLT transfusion were identified by multivariate logistic regression. Using a development data set containing 2005 to 2016 data, the Australian Cardiac Surgery Platelet Transfusion (ACSePT) risk prediction tool was developed by assigning weights to each significant predictor that corresponded to a probability of PLT transfusion. The predicted probability for each score was compared to actual PLT transfusion occurrence in a validation (2017) data set. RESULTS: The development data set contained 38 independent variables and 91 521 observations. The validation data set contained 12 529 observations. The optimal model contained 23 variables significant at P < .001 and an area under the receiver operating characteristic (ROC) curve of 0.69 (95% confidence interval [CI], 0.68-0.69). ACSePT contained nine variables and had an area under the ROC curve of 0.66 (95% CI, 0.65-0.66) and overall predicted probability of PLT transfusion of 19.8% for the validation data set compared to an observed risk of 20.3%. CONCLUSION: The ACSePT risk prediction tool is the first scoring system to predict a cardiac surgery patient's risk of receiving a PLT transfusion. It can be used to identify patients at higher risk of receiving PLT transfusions for inclusion in clinical trials and by PLT inventory managers to predict PLT demand.

Platelet storage duration and its clinical and transfusion outcomes: a systematic review.

BACKGROUND: Platelets (PLTs) are usually stored for up to 5 days prior to transfusion, although in some blood services the storage period is extended to 7 days. During storage, changes occur in both PLT and storage medium, which may lead to PLT activation and dysfunction. The clinical significance of these changes remains uncertain. METHODS: We performed a systematic review to assess the association between PLT storage time and clinical or transfusion outcomes in patients receiving allogeneic PLT transfusion. We searched studies published in English between January 2000 and July 2017 identified from MEDLINE, Embase, PubMed and the Cochrane Libraries. RESULTS: Of the 18 studies identified, five included 4719 critically ill patients (trauma, post-cardiac surgery and a heterogeneous population of critically ill patients) and 13 included 8569 haematology patients. The five studies in critically ill patients were retrospective and did not find any association between PLT storage time when PLTs were stored for up to 5 days and mortality. There was also no association between older PLTs and sepsis in the two largest studies (n = 4008 patients). Of the 13 studies in haematology patients, seven analysed prolonged storage time up to 6.5 or 7 days. Administration of fresh PLTs (less than 2 or 3 days) was associated with a significant increase in corrected count increment (CCI) compared to older PLTs in seven of the eight studies analysing this outcome. One single centre retrospective study found an increase in bleeding events in patients receiving older PLTs. CONCLUSIONS: PLT storage time does not appear to be associated with clinical outcomes, including bleeding, sepsis or mortality, in critically ill patients or haematology patients. The freshest PLTs (less than 3 days) were associated with a better CCI, although there was no impact on bleeding events, questioning the clinical significance of this association. However, there is an absence of evidence to draw definitive conclusions, especially in critically ill patients.

Duration of platelet storage and outcomes of critically ill patients.

BACKGROUND: The storage duration of platelet (PLT) units is limited to 5 to 7 days. This study investigates whether PLT storage duration is associated with patient outcomes in critically ill patients. STUDY DESIGN AND METHODS: This study was a retrospective analysis of critically ill patients admitted to the intensive care unit (ICU) of two hospitals in Australia who received one or more PLT transfusions from 2008 to 2014. Storage duration was approached in several different ways. Outcome variables were hospital mortality and ICU-acquired infection. Associations between PLT storage duration and outcomes were evaluated using multiple logistic regression and also by Cox regression. RESULTS: Among 2250 patients who received one or more PLT transfusions while in the ICU, the storage duration of PLTs was available for 64% of patients (1430). In-hospital mortality was 22.1% and ICU infection rate 7.2%. When comparing patients who received PLTs of a maximum storage duration of not more than 3, 4, or 5 days, there were no significant differences in baseline characteristics. After confounders were adjusted for, the storage duration of PLTs was not independently associated with mortality (4 days vs. ≤3 days, odds ratio [OR] 0.88, 95% confidence interval [CI] 0.59-1.30; 5 days vs. ≤3 days, OR 0.97, 95% CI 0.68-1.37) or infection (4 days vs. ≤3 days, OR 0.71, 95% CI 0.39-1.29; 5 days vs. ≤3 days, OR 1.11, 95% CI 0.67-1.83). Similar results were obtained regardless of how storage duration of PLTs was approached. CONCLUSIONS: In this large observational study in a heterogeneous ICU population, storage duration of PLTs was not associated with an increased risk of mortality or infection.

Is platelet transfusion associated with hospital-acquired infections in critically ill patients?

BACKGROUND: Platelets are commonly transfused to critically ill patients. Reports suggest an association between platelet transfusion and infection. However, there is no large study to have determined whether platelet transfusion in critically ill patients is associated with hospital-acquired infection. METHODS: We conducted a multi-centre study using prospectively maintained databases of two large academic intensive care units (ICUs) in Australia. Characteristics of patients who received platelets in ICUs between 2008 and 2014 were compared to those of patients who did not receive platelets. Association between platelet administration and infection (bacteraemia and/or bacteriuria) was modelled using multiple logistic regression and Cox regression, with blood components as time-varying covariates. A propensity covariate adjustment was also performed to verify results. RESULTS: Of the 18,965 patients included, 2250 (11.9%) received platelets in ICU with a median number of 1 platelet unit (IQR 1-3) administered. Patients who received platelets were more severely ill at ICU admission (mean Acute Physiology and Chronic Health Evaluation III score 65 (SD 29) vs 52 (SD 25), p < 0.01) and had more comorbidities (31% vs 19%, p < 0.01) than patients without platelet transfusion. Invasive mechanical ventilation (87% vs 57%, p < 0.01) and renal replacement therapy (20% vs 4%, p < 0.01) were more frequently administered in patients receiving platelets than in patients without platelets. On univariate analysis, platelet transfusion was associated with hospital-acquired infection in the ICU (7.7% vs 1.4%, p < 0.01). After adjusting for confounders, including other blood components administered, patient severity, centre, year, and diagnosis category, platelet transfusions were independently associated with infection (adjusted OR 2.56 95% CI 1.98-3.31, p < 0.001). This association was also found in survival analysis with blood components as time-varying covariates (adjusted HR 1.85, 95% CI 1.41-2.41, p < 0.001) and when only bacteraemia was considered (adjusted OR 3.30, 95% CI 2.30-4.74, p <0.001). Platelet transfusions remained associated with infection after propensity covariate adjustment. CONCLUSIONS: After adjustment for confounders, including patient severity and other blood components, platelet transfusion was independently associated with ICU-acquired infection. Further research aiming to better understand this association and to prevent this complication is warranted.

LAT1 activity of carboxylic acid bioisosteres: Evaluation of hydroxamic acids as substrates.

Large neutral amino acid transporter 1 (LAT1) is a solute carrier protein located primarily in the blood-brain barrier (BBB) that offers the potential to deliver drugs to the brain. It is also up-regulated in cancer cells, as part of a tumor's increased metabolic demands. Previously, amino acid prodrugs have been shown to be transported by LAT1. Carboxylic acid bioisosteres may afford prodrugs with an altered physicochemical and pharmacokinetic profile than those derived from natural amino acids, allowing for higher brain or tumor levels of drug and/or lower toxicity. The effect of replacing phenylalanine's carboxylic acid with a tetrazole, acylsulfonamide and hydroxamic acid (HA) bioisostere was examined. Compounds were tested for their ability to be LAT1 substrates using both cis-inhibition and trans-stimulation cell assays. As HA-Phe demonstrated weak substrate activity, its structure-activity relationship (SAR) was further explored by synthesis and testing of HA derivatives of other LAT1 amino acid substrates (i.e., Tyr, Leu, Ile, and Met). The potential for a false positive in the trans-stimulation assay caused by parent amino acid was evaluated by conducting compound stability experiments for both HA-Leu and the corresponding methyl ester derivative. We concluded that HA's are transported by LAT1. In addition, our results lend support to a recent account that amino acid esters are LAT1 substrates, and that hydrogen bonding may be as important as charge for interaction with the transporter binding site.

Metastatic Melanoma Invading the Minor Duodenal Papilla in a Patient With Pancreas Divisum Causing Acute Recurrent Pancreatitis: A Case Report.

Metastasis to the gastrointestinal (GI) tract should always be a consideration when melanoma, particularly metastatic disease, is diagnosed. While metastasis to the small intestine is common, given its rich blood supply, metastasis to the pancreatic ducts is extremely rare. In patients with pancreatic divisum, disease spread to the minor papilla can greatly increase the chance of developing pancreatitis due to the potential for increased pancreatic intraductal pressure. We present one unique case of metastatic melanoma to the minor duodenal papilla causing pancreatitis.

Transfusion practices in intensive care units: An Australian and New Zealand point prevalence study.

Objective: To describe current transfusion practices in intensive care units (ICUs) in Australia and New Zealand, compare them against national guidelines, and describe how viscoelastic haemostatic assays (VHAs) are used in guiding transfusion decisions. Design setting and participants: Prospective, multicentre, binational point-prevalence study. All adult patients admitted to participating ICUs on a single day in 2021. Main outcome measures: Transfusion types, amounts, clinical reasons, and triggers; use of anti-platelet medications, anti-coagulation, and VHA. Results: Of 712 adult patients in 51 ICUs, 71 (10%) patients received a transfusion during the 24hr period of observation. Compared to patients not transfused, these patients had higher Acute Physiology and Chronic Health Evaluation II scores (19 versus 17, p = 0.02), a greater proportion were mechanically ventilated (49.3% versus 37.3%, p < 0.05), and more had systemic inflammatory response syndrome (70.4% versus 51.3%, p < 0.01). Overall, 63 (8.8%) patients received red blood cell (RBC) transfusions, 10 (1.4%) patients received platelet transfusions, 6 (0.8%) patients received fresh frozen plasma (FFP), and 5 (0.7%) patients received cryoprecipitate. VHA was available in 42 (82.4%) sites but only used in 6.6% of transfusion episodes when available. Alignment with guidelines was found for 98.6% of RBC transfusions, but only 61.6% for platelet, 28.6% for FFP, and 20% for cryoprecipitate transfusions. Conclusions: Non-RBC transfusion decisions are often not aligned with guidelines and VHA is commonly available but rarely used to guide transfusions. Better evidence to guide transfusions in ICUs is needed.

Resident tissue-specific mesenchymal progenitor cells contribute to fibrogenesis in human lung allografts.

Fibrotic obliteration of the small airways leading to progressive airflow obstruction, termed bronchiolitis obliterans syndrome (BOS), is the major cause of poor outcomes after lung transplantation. We recently demonstrated that a donor-derived population of multipotent mesenchymal stem cells (MSCs) can be isolated from the bronchoalveolar lavage (BAL) fluid of human lung transplant recipients. Herein, we study the organ specificity of these cells and investigate the role of local mesenchymal progenitors in fibrogenesis after lung transplantation. We demonstrate that human lung allograft-derived MSCs uniquely express embryonic lung mesenchyme-associated transcription factors with a 35,000-fold higher expression of forkhead/winged helix transcription factor forkhead box (FOXF1) noted in lung compared with bone marrow MSCs. Fibrotic differentiation of MSCs isolated from normal lung allografts was noted in the presence of profibrotic mediators associated with BOS, including transforming growth factor-ß and IL-13. MSCs isolated from patients with BOS demonstrated increased expression of α-SMA and collagen I when compared with non-BOS controls, consistent with a stable in vivo fibrotic phenotype. FOXF1 mRNA expression in the BAL cell pellet correlated with the number of MSCs in the BAL fluid, and myofibroblasts present in the fibrotic lesions expressed FOXF1 by in situ hybridization. These data suggest a key role for local tissue-specific, organ-resident, mesenchymal precursors in the fibrogenic processes in human adult lungs.

Mesenchymal stromal cells in bronchoalveolar lavage as predictors of bronchiolitis obliterans syndrome.

RATIONALE: Bronchoalveolar lavage fluid (BAL) from human lung allografts demonstrates the presence of a multipotent mesenchymal stromal cell population. However, the clinical relevance of this novel cellular component of BAL and its association with bronchiolitis obliterans syndrome (BOS), a disease marked by progressive airflow limitation secondary to fibrotic obliteration of the small airways, remains to be determined. OBJECTIVES: In this study we investigate the association of number of mesenchymal stromal cells in BAL with development of BOS in human lung transplant recipients. METHODS: Mesenchymal colony-forming units (CFUs) were quantitated in a cohort of 405 BAL samples obtained from 162 lung transplant recipients. Poisson generalized estimating equations were used to determine the predictors of BAL mesenchymal CFU count. MEASUREMENTS AND MAIN RESULTS: Higher CFU counts were noted early post-transplantation; time from transplant to BAL of greater than 3 months predicted 0.4-fold lower CFU counts (P = 0.0001). BOS diagnosis less than or equal to 365 days before BAL was associated with a 2.11-fold higher CFU count (P = 0.02). There were 2.62- and 2.70-fold higher CFU counts noted in the presence of histologic diagnosis of bronchiolitis obliterans (P = 0.05) and organizing pneumonia (0.0003), respectively. In BAL samples obtained from BOS-free patients greater than 6 months post-transplantation (n = 173), higher mesenchymal CFU counts (≥10) significantly predicted BOS onset in both univariate (hazard ratio, 5.61; 95% CI, 3.03-10.38; P < 0.0001) and multivariate (hazard ratio, 5.02; 95% CI, 2.40-10.51; P < 0.0001) Cox regression analysis. CONCLUSIONS: Measurement of mesenchymal CFUs in the BAL provides predictive information regarding future BOS onset.

Pigmented orbital mass due to remote pencil trauma.

A 45-year-old woman with a medial canthal mass associated with new-onset epiphora was found to have a darkly pigmented mass arising from the lacrimal sac wall because of intranasal pencil trauma 40 years before. Resection of the graphite-induced granuloma with dacryocystorhinostomy was curative.

A Case of Undifferentiated Pleomorphic Sarcoma.

Suspicion for soft tissue malignancy of the hand is usually low because most tumors of the hand are small and benign. We present a case of an elderly female who presented with a rapidly enlarging, ulcerating hand mass over a few months. She was diagnosed with undifferentiated pleomorphic sarcoma (UPS), a high-grade, aggressive soft-tissue sarcoma. Computed tomography (CT) of the chest was conducted for staging purposes. It showed multiple subcentimeter pulmonary nodules, findings that were initially worrisome for metastatic disease but later proved to be newly and incidentally diagnosed granulomatous disease of the lungs. This case highlights the importance of early recognition of potential malignancy in soft-tissue tumors of the hand to facilitate proper referral and initiation of appropriate oncologic therapies. Due to early diagnosis and intervention, our patient had locally advanced disease without metastasis, a type of cancer known to have a high degree of metastatic potential.

Partial deletions of the autoregulatory C-terminal domain of Artemis and their effect on its nuclease activity.

Artemis is a 692 aa nuclease that is essential for opening hairpins during vertebrate V(D)J recombination. Artemis is also important in the DNA repair of double-strand breaks via the nonhomologous DNA end joining (NHEJ) pathway. Therefore, absence of Artemis has been shown to result not only in the blockage of lymphocyte development in vertebrates, but also sensitivity of organisms and cells to double-strand break-inducing events that arise in the course of normal metabolism. Nonhomologous DNA end joining (NHEJ) is the major pathway for the repair of double-strand DNA breaks in most vertebrate cells during most of the cell cycle, including in resting cells. Artemis is the primary nuclease for resection of damaged DNA at double-strand breaks. Artemis alone is inactive as an endonuclease, though it has 5'-exonuclease activity. The endonuclease activity requires physical interaction with DNA-PKcs and subsequent activation steps. Truncation of the C-terminal half of Artemis permits Artemis to be active, even without DNA-PKcs. Here we create a systematic set of deletions from the Artemis C-terminus to determine the minimal extent of C-terminal deletion for Artemis to function in a DNA-PKcs-independent manner. We discuss these data in the context of recent structural studies. The results will be useful in future studies to determine the full range of functions of the C-terminal region of Artemis in the regulation of its endonuclease activity.

Epithelial interactions and local engraftment of lung-resident mesenchymal stem cells.

Multipotent mesenchymal progenitor cells, termed "mesenchymal stem cells" (MSCs), have been demonstrated to reside in human adult lungs. However, there is little information regarding the associations of these local mesenchymal progenitors with other resident somatic cells and their potential for therapeutic use. Here we provide in vivo and in vitro evidence for the ability of human adult lung-resident MSCs (LR-MSCs) to interact with the local epithelial cells. The in vivo retention and localization of human LR-MSCs in an alveolar microenvironment was investigated by placing PKH-26 or DsRed lentivirus-labeled human LR-MSCs in the lungs of immunodeficient (SCID) mice. At 3 weeks after intratracheal administration, 19.3 ± 3.21% of LR-MSCs were recovered, compared with 3.47 ± 0.51% of control fibroblasts, as determined by flow cytometry. LR-MSCs were found to persist in murine lungs for up to 6 months and demonstrated preferential localization to the corners of the alveoli in close proximity to type II alveolar epithelial cells, the progenitor cells of the alveolar epithelium. In vitro, LR-MSCs established gap junction communications with lung alveolar and bronchial epithelial cells and demonstrated an ability to secrete keratinocyte growth factor, an important modulator of epithelial cell proliferation and differentiation. Gap junction communications were also demonstrable between LR-MSCs and resident murine cells in vivo. This study demonstrates, for the first time, an ability of tissue-specific MSCs to engraft in their organ of origin and establishes a pathway of bidirectional interaction between these mesenchymal progenitors and adult somatic epithelial cells in the lung.

Epidemiology of Massive Transfusion - A Common Intervention in Need of a Definition.

While massive transfusion (MT) recipients account for a small proportion of all transfused patients, they account for approximately 10% of blood products issued. Furthermore, MT events pose organizational and logistical challenges for health care providers, laboratory and transfusion services. Overall, the majority of MT events are to support major bleeding in surgical patients, trauma and gastrointestinal hemorrhage. The clinical context in which the bleeding event occurred, the number of blood products required, patient age and comorbidities are the most important predictors of outcomes for short- and long-term survival. These data are important to inform blood services, clinicians and health care providers in order to improve care and outcomes for patients with major bleeding. There is no standard accepted definition of MT, with most definitions based on number of blood components administered within a certain time-period or activation of MT protocol. The type of definition used has implications for the clinical characteristics of MT recipients included in epidemiological and interventional studies. In order to understand trends in incidence of MT, variation in blood utilization and patient outcomes, and to harmonize research outcomes, a standard and universally accepted definition of MT is urgently required.

Adrenal Infarction in Pregnancy Secondary to Elevated Plasma Factor VIII Activity.

Unilateral adrenal infarction is a rare cause of acute abdomen in pregnancy (AAP). Its presentation is non-specific and requires a high index of suspicion with a low threshold to obtain radiographic imaging for diagnosis. Evaluating AAP is challenging as diagnostic radiographic imaging is often limited in relation to radiation exposure to the developing fetus. We describe a case of a 24-year-old pregnant female who presented with severe acute abdominal pain. The patient's pain was refractory to intravenous analgesics and ultrasonography was inconclusive. Computed tomography (CT) scan was not obtained due to the risk of radiation exposure to the developing fetus. Due to the persistence of pain and suspicions for other serious etiologies, magnetic resonance imaging (MRI) was completed and the patient was diagnosed with acute unilateral adrenal infarction. In this case report, unilateral adrenal infarction was likely secondary to elevated plasma factor VIII levels. Even with the physiological elevation of factor VIII levels during pregnancy, levels greater than 150 IU/dL confer greater than five-fold increased risk of venous thrombosis. Once hemorrhage is excluded, patients should be started on therapeutic anticoagulation to prevent progression of adrenal infarct or infarction of the contralateral adrenal gland. Prompt recognition and treatment of acute adrenal infarction during pregnancy are of paramount importance to prevent adverse outcomes for both the mother and fetus.

Structural insights into the function of the catalytically active human Taspase1.

Taspase1 is an Ntn-hydrolase overexpressed in primary human cancers, coordinating cancer cell proliferation, invasion, and metastasis. Loss of Taspase1 activity disrupts proliferation of human cancer cells in vitro and in mouse models of glioblastoma. Taspase1 is synthesized as an inactive proenzyme, becoming active upon intramolecular cleavage. The activation process changes the conformation of a long fragment at the C-terminus of the α subunit, for which no full-length structural information exists and whose function is poorly understood. We present a cloning strategy to generate a circularly permuted form of Taspase1 to determine the crystallographic structure of active Taspase1. We discovered that this region forms a long helix and is indispensable for the catalytic activity of Taspase1. Our study highlights the importance of this element for the enzymatic activity of Ntn-hydrolases, suggesting that it could be a potential target for the design of inhibitors with potential to be developed into anticancer therapeutics.

Evidence for tissue-resident mesenchymal stem cells in human adult lung from studies of transplanted allografts.

The origin and turnover of connective tissue cells in adult human organs, including the lung, are not well understood. Here, studies of cells derived from human lung allografts demonstrate the presence of a multipotent mesenchymal cell population, which is locally resident in the human adult lung and has extended life span in vivo. Examination of plastic-adherent cell populations in bronchoalveolar lavage samples obtained from 76 human lung transplant recipients revealed clonal proliferation of fibroblast-like cells in 62% (106 of 172) of samples. Immunophenotyping of these isolated cells demonstrated expression of vimentin and prolyl-4-hydroxylase, indicating a mesenchymal phenotype. Multiparametric flow cytometric analyses revealed expression of cell-surface proteins, CD73, CD90, and CD105, commonly found on mesenchymal stem cells (MSCs). Hematopoietic lineage markers CD14, CD34, and CD45 were absent. Multipotency of these cells was demonstrated by their capacity to differentiate into adipocytes, chondrocytes, and osteocytes. Cytogenetic analysis of cells from 7 sex-mismatched lung transplant recipients harvested up to 11 years after transplant revealed that 97.2% +/- 2.1% expressed the sex genotype of the donor. The presence of MSCs of donor sex identity in lung allografts even years after transplantation provides what we believe to be the first evidence for connective tissue cell progenitors that reside locally within a postnatal, nonhematopoietic organ.