A systematic investigation of production of synthetic prions from recombinant prion protein

According to the protein-only hypothesis, infectious mammalian prions, which exist as distinct strains with discrete biological properties, consist of multichain assemblies of misfolded cellular prion protein (PrP). A critical test would be to produce prion strains synthetically from defined components. Crucially, high-titre 'synthetic' prions could then be used to determine the structural basis of infectivity and strain diversity at the atomic level. While there have been multiple reports of production of prions from bacterially expressed recombinant PrP using various methods, systematic production of high-titre material in a form suitable for structural analysis remains a key goal. Here, we report a novel high-throughput strategy for exploring a matrix of conditions, additives and potential cofactors that might generate high-titre prions from recombinant mouse PrP, with screening for infectivity using a sensitive automated cell-based bioassay. Overall, approximately 20 000 unique conditions were examined. While some resulted in apparently infected cell cultures, this was transient and not reproducible. We also adapted published methods that reported production of synthetic prions from recombinant hamster PrP, but again did not find evidence of significant infectious titre when using recombinant mouse PrP as substrate. Collectively, our findings are consistent with the formation of prion infectivity from recombinant mouse PrP being a rare stochastic event and we conclude that systematic generation of prions from recombinant PrP may only become possible once the detailed structure of authentic ex vivo prions is solved

Long-term beneficial effect of islet transplantation on diabetic macro-/microangiopathy in type 1 diabetic kidney-transplanted patients

OBJECTIVE: Our aim was to evaluate the long-term effects of transplanted islets on diabetic macro-/microangiopathy in type 1 diabetic kidney-transplanted patients. RESEARCH DESIGN AND METHODS: A total of 34 type 1 diabetic kidney-transplanted patients underwent islet transplantation and were divided into two groups: successful islet-kidney transplantation (SI-K; 21 patients, fasting C-peptide serum concentration >0.5 ng/ml for >1 year) and unsuccessful islet-kidney transplantation (UI-K; 13 patients, fasting C-peptide serum concentration <0.5 ng/ml). Patients cumulative survival, cardiovascular death rate, and atherosclerosis progression were compared in the two groups. Skin biopsies, endothelial dependent dilation (EDD), nitric oxide (NO) levels, and atherothrombotic risk factors [von Willebrand factor (vWF) and D-dimer fragment (DDF)] were studied cross-sectionally. RESULTS: The SI-K group showed a significant better patient survival rate (SI-K 100, 100, and 90% vs. UI-K 84, 74, and 51% at 1, 4, and 7 years, respectively, P = 0.04), lower cardiovascular death rate (SI-K 1/21 vs. UI-K 4/13, chi(2) = 3.9, P = 0.04), and lower intima-media thickness progression than the UI-K group (SI-K group: delta1-3 years -13 +/- 30 micro m vs. UI-K group: delta1-3 years 245 +/- 20 micro m, P = 0.03) with decreased signs of endothelial injuring at skin biopsy. Furthermore, the SI-K group showed a higher EDD than the UI-K group (EDD: SI-K 7.8 +/- 4.5% vs. UI-K 0.5 +/- 2.7%, P = 0.02), higher basal NO (SI-K 42.9 +/- 6.5 vs. UI-K 20.2 +/- 6.8 micro mol/l, P = 0.02), and lower levels of vWF (SI-K 138.6 +/- 15.3 vs. UI-K 180.6 +/- 7.0%, P = 0.02) and DDF (SI-K 0.61 +/- 0.22 vs. UI-K 3.07 +/- 0.68 micro g/ml, P < 0.01). C-peptide-to-creatinine ratio correlated positively with EDD and NO and negatively with vWF and DDF. CONCLUSIONS: Successful islet transplantation improves survival, cardiovascular, and endothelial function in type 1 diabetic kidney-transplanted patient

Ovarian remnant syndrome after laparoscopic ovariectomy in a cat

The first case of accidental implantation of ovarian tissue at the port site after laparoscopic ovariectomy in a cat was described. A 2-year-old, spayed female, domestic cat showed recurrent behavioral oestrus signs (vocalization, increased playfulness, and lordosis) four months after laparoscopic ovariectomy. Abdominal ultrasound revealed bilateral endometrial hyperplasia and an inhomogeneous mass attached to the abdominal wall at the port site of previous laparoscopic ovariectomy. Ovarian remnant syndrome was supposed. Diagnostic laparoscopy confirmed the presence of a mass that was 2 cm in diameter and macroscopically similar to ovarian tissue. Laparoscopy was converted to laparotomy to remove the abnormal tissue and the uterus. Histopathological findings showed follicles of various maturation stages in the mass, and confirmed endometrial hyperplasia. Histology underscored that ovarian remnant tissue resumed function. No clinical signs consistent with ovarian remnant syndrome were reported six months after removal of the abnormal tissue. In conclusion, removal of the ovaries through the laparoscopic port site may cause fortuitous ovarian tissue implantation at the abdominal wall in the cat

Metformin increases skeletal muscle lactate production in pigs: a microdialysis study

Introduction Lactic acidosis during metformin intoxication is mainly attributed to impaired hepatic lactate clearance [1]. The aim of this present work was to clarify whether metformin at high dose also increases skeletal muscle lactate production. Methods Reverse microdialysis was used in six healthy, sedated and mechanically ventilated pigs, equipped with two skeletal muscle catheters (CMA Microdialysis AB, Sweden). Following a baseline recording, a continuous infusion of saline (control) or metformin diluted in saline (1 mol/l) began. Outfl ow lactate concentration was measured every 3 hours, up to 12 hours. Results Data are presented as the mean and standard deviation in Figure 1. The interaction between infusion (saline vs. metformin) and time was statistically signifi cant (P = 0.02; two-way repeated-measures ANOVA). Conclusions In skeletal muscle, a high dose of metformin increases interstitial lactate levels, a fi nding consistent with local lactate overproduction. Reference 1. Lalau JD: Drug Saf 2010, 33:727-740

Case Report: Late-Onset Congenital Adrenal Hyperplasia and Acute Covid-19 Infection in a Pregnant Woman: Multidisciplinary Management

Background The impact of the Covid-19 infection on patients with chronic endocrine disease is not fully known. We describe here the first case of a pregnant woman with Covid-19 acute infection and non-classical congenital adrenal hyperplasia (NCAH). Case description A woman at 36 weeks of gestation was referred to our Maternity Hospital for premature rupture of membranes (PROM). Her medical history was positive for NCAH on chronic steroid replacement till the age of 17 years (cortisone acetate and dexamethasone, both in the morning). At admission, her naso-oro-pharyngeal swab resulted positive for SARS-CoV-2. Due to hyperpyrexia and late preterm PROM, cesarean section was planned, and she was started on a 100 mg-bolus of hydrocortisone, followed by continuous infusion of 200 mg/24 h. A female neonate in good clinical condition and with a negative nasopharyngeal Covid-19 swab was delivered. On second postpartum day, the mother was in good condition and was switched to oral steroid therapy. On third postpartum day she worsened, with radiological signs of acute pulmonary embolism. Oro-tracheal intubation and mechanical ventilation were started, and she was switched back to intravenous steroid therapy. On April 30, pulmonary embolism was resolved, and on May 13th she was discharged in good condition. Conclusions We report the first case of Covid-19 acute infection that occurred in late-pregnancy in a woman with NCAH on chronic steroid replacement. The management of the patient in a reference center with early involvement of a multidisciplinary team granted prompt care and adequate protection for all the involved sanitary operators

Cortical microstructural changes predict tau accumulation and episodic memory decline in older adults harboring amyloid

Non-invasive diffusion-weighted imaging (DWI) to assess brain microstructural changes via cortical mean diffusivity (cMD) has been shown to be cross-sectionally associated with tau in cognitively normal older adults, suggesting that it might be an early marker of neuronal injury. Here, we investigated how regional cortical microstructural changes measured by cMD are related to the longitudinal accumulation of regional tau as well as to episodic memory decline in cognitively normal individuals harboring amyloid pathology. 122 cognitively normal participants from the Harvard Aging Brain Study underwent DWI, T1w-MRI, amyloid and tau PET imaging, and Logical Memory Delayed Recall (LMDR) assessments. We assessed whether the interaction of baseline amyloid status and cMD (in entorhinal and inferior-temporal cortices) was associated with longitudinal regional tau accumulation and with longitudinal LMDR using separate linear mixed-effects models. We find a significant interaction effect of the amyloid status and baseline cMD in predicting longitudinal tau in the entorhinal cortex (p = 0.044) but not the inferior temporal lobe, such that greater baseline cMD values predicts the accumulation of entorhinal tau in amyloid-positive participants. Moreover, we find a significant interaction effect of the amyloid status and baseline cMD in the entorhinal cortex (but not inferior temporal cMD) in predicting longitudinal LMDR (p < 0.001), such that baseline entorhinal cMD predicts the episodic memory decline in amyloid-positive participants. The combination of amyloidosis and elevated cMD in the entorhinal cortex may help identify individuals at short-term risk of tau accumulation and Alzheimer's Disease-related episodic memory decline, suggesting utility in clinical trials

Epidural Corticosteroids, Lumbar Spinal Drainage, and Selective Hemodynamic Control for the Prevention of Spinal Cord Ischemia in Thoracoabdominal Endovascular Aortic Repair: A New Clinical Protocol

Introduction: In patients undergoing thoracoabdominal aorta repair, spinal cord ischemia (SCI) remains one of the most common and important complications resulting in transient paraparesis through to permanent flaccid paraplegia. In this manuscript, after a brief introduction to spinal cord ischemia complication and its prevention in thoracoabdominal endovascular aortic repair (TEVAR), we propose a new clinical protocol&nbsp;potentially able to prevent such complication. Methods: The proposed protocol suggests the use of high dosages of corticosteroids by epidural route, along with drainage of cerebrospinal fluid and controlled vascular hypertension, to reduce the incidence of SCI in TEVAR. Moreover, we paid particular attention to the control of the hemodynamic parameters to obtain adequate peripheral tissue perfusion (oxygen delivery), including in the spinal cord. Results: We applied this new protocol in 50 consecutive patients treated with TEVAR for thoracoabdominal aortic aneurysms (TAAs); 47 patients completed the procedure: 27 patients Crawford type&nbsp;I and 20 Crawford type&nbsp;II. Three patients died during surgery because of untreatable aneurysm rupture. The results show that in all patients there were no cases of SCI, after 5&nbsp;days from TEVAR. Discussion: To the best of our knowledge, there are no clinical studies on the use of epidural corticosteroids in patients undergoing treatment of aortic syndrome (both in "open surgery" and endovascular aortic repair). This initial study on 50 consecutive patients has shown that the clinical protocol used could be of great interest to prevent one of the worse complications of TEVAR. Its limitations are the low number of patients studied till now, and the non-randomized protocol adopted. Further studies would be necessary. Conclusion: Our experience and the results obtained with this new perioperative protocol with epidural corticosteroid and accurate hemodynamic control have been encouraging and it seems a valid proposal to be explored in future by well-structured prospective, randomized protocols.Introduction: In patients undergoing thoracoabdominal aorta repair, spinal cord ischemia (SCI) remains one of the most common and important complications resulting in transient paraparesis through to permanent flaccid paraplegia. In this manuscript, after a brief introduction to spinal cord ischemia complication and its prevention in thoracoabdominal endovascular aortic repair (TEVAR), we propose a new clinical protocol&nbsp;potentially able to prevent such complication. Methods: The proposed protocol suggests the use of high dosages of corticosteroids by epidural route, along with drainage of cerebrospinal fluid and controlled vascular hypertension, to reduce the incidence of SCI in TEVAR. Moreover, we paid particular attention to the control of the hemodynamic parameters to obtain adequate peripheral tissue perfusion (oxygen delivery), including in the spinal cord. Results: We applied this new protocol in 50 consecutive patients treated with TEVAR for thoracoabdominal aortic aneurysms (TAAs); 47 patients completed the procedure: 27 patients Crawford type&nbsp;I and 20 Crawford type&nbsp;II. Three patients died during surgery because of untreatable aneurysm rupture. The results show that in all patients there were no cases of SCI, after 5&nbsp;days from TEVAR. Discussion: To the best of our knowledge, there are no clinical studies on the use of epidural corticosteroids in patients undergoing treatment of aortic syndrome (both in “open surgery” and endovascular aortic repair). This initial study on 50 consecutive patients has shown that the clinical protocol used could be of great interest to prevent one of the worse complications of TEVAR. Its limitations are the low number of patients studied till now, and the non-randomized protocol adopted. Further studies would be necessary. Conclusion: Our experience and the results obtained with this new perioperative protocol with epidural corticosteroid and accurate hemodynamic control have been encouraging and it seems a valid proposal to be explored in future by well-structured prospective, randomized protocols

Time course of risk factors associated with mortality of 1260 critically ill patients with COVID-19 admitted to 24 Italian intensive care units

94noopenPurpose: To evaluate the daily values and trends over time of relevant clinical, ventilatory and laboratory parameters during the intensive care unit (ICU) stay and their association with outcome in critically ill patients with coronavirus disease 19 (COVID-19). Methods: In this retrospective–prospective multicentric study, we enrolled COVID-19 patients admitted to Italian ICUs from February 22 to May 31, 2020. Clinical data were daily recorded. The time course of 18 clinical parameters was evaluated by a polynomial maximum likelihood multilevel linear regression model, while a full joint modeling was fit to study the association with ICU outcome. Results: 1260 consecutive critically ill patients with COVID-19 admitted in 24 ICUs were enrolled. 78% were male with a median age of 63 [55–69] years. At ICU admission, the median ratio of arterial oxygen partial pressure to fractional inspired oxygen (PaO2/FiO2) was 122 [89–175] mmHg. 79% of patients underwent invasive mechanical ventilation. The overall mortality was 34%. Both the daily values and trends of respiratory system compliance, PaO2/FiO2, driving pressure, arterial carbon dioxide partial pressure, creatinine, C-reactive protein, ferritin, neutrophil, neutrophil–lymphocyte ratio, and platelets were associated with survival, while for lactate, pH, bilirubin, lymphocyte, and urea only the daily values were associated with survival. The trends of PaO2/FiO2, respiratory system compliance, driving pressure, creatinine, ferritin, and C-reactive protein showed a higher association with survival compared to the daily values. Conclusion: Daily values or trends over time of parameters associated with acute organ dysfunction, acid–base derangement, coagulation impairment, or systemic inflammation were associated with patient survival.openZanella A.; Florio G.; Antonelli M.; Bellani G.; Berselli A.; Bove T.; Cabrini L.; Carlesso E.; Castelli G.P.; Cecconi M.; Citerio G.; Coloretti I.; Corti D.; Dalla Corte F.; De Robertis E.; Foti G.; Fumagalli R.; Girardis M.; Giudici R.; Guiotto L.; Langer T.; Mirabella L.; Pasero D.; Protti A.; Ranieri M.V.; Rona R.; Scudeller L.; Severgnini P.; Spadaro S.; Stocchetti N.; Vigano M.; Pesenti A.; Grasselli G.; Aspesi M.; Baccanelli F.; Bassi F.; Bet A.; Biagioni E.; Biondo A.; Bonenti C.; Bottino N.; Brazzi L.; Buquicchio I.; Busani S.; Calini A.; Calligaro P.; Cantatore L.P.; Carelli S.; Carsetti A.; Cavallini S.; Cimicchi G.; Coppadoro A.; Dall'Ara L.; Di Gravio V.; Erba M.; Evasi G.; Facchini A.; Fanelli V.; Feliciotti G.; Fusarini C.F.; Ferraro G.; Gagliardi G.; Garberi R.; Gay H.; Giacche L.; Grieco D.; Guzzardella A.; Longhini F.; Manzan A.; Maraggia D.; Milani A.; Mischi A.; Montalto C.; Mormina S.; Noseda V.; Paleari C.; Pedeferri M.; Pezzi A.; Pizzilli G.; Pozzi M.; Properzi P.; Rauseo M.; Russotto V.; Saccarelli L.; Servillo G.; Spano S.; Tagliabue P.; Tonetti T.; Tullo L.; Vetrugno L.; Vivona L.; Volta C.A.; Zambelli V.; Zanoni A.Zanella, A.; Florio, G.; Antonelli, M.; Bellani, G.; Berselli, A.; Bove, T.; Cabrini, L.; Carlesso, E.; Castelli, G. P.; Cecconi, M.; Citerio, G.; Coloretti, I.; Corti, D.; Dalla Corte, F.; De Robertis, E.; Foti, G.; Fumagalli, R.; Girardis, M.; Giudici, R.; Guiotto, L.; Langer, T.; Mirabella, L.; Pasero, D.; Protti, A.; Ranieri, M. V.; Rona, R.; Scudeller, L.; Severgnini, P.; Spadaro, S.; Stocchetti, N.; Vigano, M.; Pesenti, A.; Grasselli, G.; Aspesi, M.; Baccanelli, F.; Bassi, F.; Bet, A.; Biagioni, E.; Biondo, A.; Bonenti, C.; Bottino, N.; Brazzi, L.; Buquicchio, I.; Busani, S.; Calini, A.; Calligaro, P.; Cantatore, L. P.; Carelli, S.; Carsetti, A.; Cavallini, S.; Cimicchi, G.; Coppadoro, A.; Dall'Ara, L.; Di Gravio, V.; Erba, M.; Evasi, G.; Facchini, A.; Fanelli, V.; Feliciotti, G.; Fusarini, C. F.; Ferraro, G.; Gagliardi, G.; Garberi, R.; Gay, H.; Giacche, L.; Grieco, D.; Guzzardella, A.; Longhini, F.; Manzan, A.; Maraggia, D.; Milani, A.; Mischi, A.; Montalto, C.; Mormina, S.; Noseda, V.; Paleari, C.; Pedeferri, M.; Pezzi, A.; Pizzilli, G.; Pozzi, M.; Properzi, P.; Rauseo, M.; Russotto, V.; Saccarelli, L.; Servillo, G.; Spano, S.; Tagliabue, P.; Tonetti, T.; Tullo, L.; Vetrugno, L.; Vivona, L.; Volta, C. A.; Zambelli, V.; Zanoni, A