Mitochondrial complex I activity in microglia sustains neuroinflammation.

Sustained smouldering, or low-grade activation, of myeloid cells is a common hallmark of several chronic neurological diseases, including multiple sclerosis1. Distinct metabolic and mitochondrial features guide the activation and the diverse functional states of myeloid cells2. However, how these metabolic features act to perpetuate inflammation of the central nervous system is unclear. Here, using a multiomics approach, we identify a molecular signature that sustains the activation of microglia through mitochondrial complex I activity driving reverse electron transport and the production of reactive oxygen species. Mechanistically, blocking complex I in pro-inflammatory microglia protects the central nervous system against neurotoxic damage and improves functional outcomes in an animal disease model in vivo. Complex I activity in microglia is a potential therapeutic target to foster neuroprotection in chronic inflammatory disorders of the central nervous system3.

High prevalence of long-term olfactory, gustatory, and chemesthesis dysfunction in post-COVID-19 patients: a matched case-control study with one-year follow-up using a comprehensive psychophysical evaluation.

BACKGROUND: Using an age and gender matched-pair case-control study, we aimed to estimate the long-term prevalence of psychophysical olfactory, gustatory , and chemesthesis impairment at least one year after SARS-CoV-2 infection considering the background of chemosensory dysfunction in non-COVID-19 population. METHODOLOGY: This case-controlled study included 100 patients who were home-isolated for mildly symptomatic COVID-19 between March and April 2020. One control regularly tested for SARS-CoV-2 infection and always tested negative was matched to each case according to gender and age. Chemosensory function was investigated by a comprehensive psychophysical evaluation including ortho- and retronasal olfaction and an extensive assessment of gustatory function. Differences in chemosensory parameters were evaluated through either Fisher’s exact test or Kruskal-Wallis test. RESULTS: The psychophysical assessment of chemosensory function took place after a median of 401 days from the first SARS-CoV-2 positive swab. The evaluation of orthonasal smell identified 46% and 10% of cases and controls, respectively, having olfactory dysfunction, with 7% of COVID-19 cases being functionally anosmic. Testing of gustatory function revealed a 27% of cases versus 10% of controls showing a gustatory impairment. Nasal trigeminal sensitivity was significantly lower in cases compared to controls. Persistent chemosensory impairment was associated with emotional distress and depression. CONCLUSION: More than one year after the onset of COVID-19, cases exhibited an excess of olfactory, gustatory , and chemesthesis disturbances compared to matched-pair controls with these symptoms being associated to emotional distress and depression.

Trans-stomal single-port laparoscopic Hartmann's reversal is an efficacious and efficient procedure: a case-controlled study.

BACKGROUND: Hartman's reversal remains challenging and is associated with a widely variable success rate. In a previous study, we reported that laparoscopy may lower the mortality and morbidity rates of the procedure. The aim of the current study was to assess the operative results of single-port laparoscopic Hartmann's reversal (SP-HR) as compared to the more standard, multi-port laparoscopic variant (MP-HR). METHODS: We performed a retrospective, non-randomized, case-controlled study of 44 consecutive patients who had SP-HR (Group A) compared to 44 patients who had MP-HR (Group B). The study was conducted in a high-volume colorectal unit in a 1200-bed university affiliated hospital, The Poissy-Saint Germain Medical Complex, France. RESULTS: Preoperative patients' characteristics (sex, body mass index, American Society of Anesthesiologists status, prior surgery, comorbidities, colonic disease) were comparable in both groups. The conversion rate was 13.6% and 4.5% in Group A and in Group B, respectively (p = 0.084) and consisted of placement of any additional ports. Conversion to open surgery did not occur in any patient in either group (p = 1). Mean operative time was shorter in Group A than in in Group B, (105 vs. 155 min; p = 0.0133). The mortality rate was 2.2% in Group A and 0% in Group B (p = 0.3145). The overall morbidity rate was 11.4% in Group A and 18.2% in Group B (p = 0.5344). The median length of hospital stay was significantly shorter in Group than in Group B (4.8 vs. 6.8 days; p = 0.0102). CONCLUSIONS: The SP-HR technique was found to be safe and efficient. It compares favorably with MP-HR. Moreover, indirect cost savings could be induced by the reduction in the length of hospital stay.

Production of cheese from donkey milk as influenced by addition of transglutaminase.

Donkey milk is characterized by low contents of total solids, fat, and caseins, especially κ-casein, which results in formation of a very weak gel upon renneting. The objective of this study was to evaluate the effect of fortification of donkey milk with microbial transglutaminase (MTGase) for cheesemaking in relation to different enzyme addition protocols (patterns, PAT). Four independent trials were performed using MTGase (5.0 U/g of milk protein) according to the following experimental patterns: control (no MTGase addition); MTGase addition (40°C) 15 min before starter inoculation (PAT1); addition of MTGase to milk simultaneously with starter culture (40°C) (PAT2); and MTGase addition simultaneously with rennet (42°C) in acidified milk (pH 6.3) (PAT3). Evolution of pH during acidification, cheesemaking parameters, and proximal composition and color of cheese at 24 h were recorded. The protein fractions of cheese and whey were investigated by urea-PAGE and sodium dodecyl sulfate-PAGE. Addition of MTGase had no significant effect on moisture, protein, fat, or cheese yield. The addition of MTGase with rennet (PAT3) improved curd firmness compared with the control. Among the different patterns of MTGase addition, PAT3 reduced gel formation time, time between rennet addition and cheese molding, and weight loss of cheese at 24 h. The PAT3 treatment also resulted in the lowest lightness and highest yellowness color values of the cheese. Sodium dodecyl sulfate-PAGE of cheeses revealed that MTGase modified the protein pattern in the high-molecular-weight zone (range 37-75 kDa) compared with the control. Of the MTGase protocols, PAT3 showed better casein retention in cheese, as confirmed by the lanes of α- and ß-caseins in the electropherogram of the whey, which was subtler for this protocol. In conclusion, MTGase may be used in cheese production from donkey milk to improve curd firmness; MTGase should be added simultaneously with the rennet.

Preliminary experience with umbilical stoma in transumbilical single-port colorectal surgery.

BACKGROUND: The umbilicus, an embryological natural orifice, is increasingly used as the only access route during single-incision laparoscopic surgery (SILS) for colorectal disease. As a part of some of these procedures, a temporary, diverting ostomy could be exteriorized through the umbilicus itself. Theoretical advantages include better preservation of the abdominal wall and potentially superior cosmetic results. The aim of the present study was to evaluate our preliminary experience in SILS colorectal resection with umbilical stoma (u-stoma). METHODS: We retrospectively reviewed all colorectal patients operated using SILS for benign or malignant disease at Paris Poissy Medical Center. Patients were selected for consideration of u-stoma with our stoma therapists. RESULTS: Between January 2010 and December 2016, 234 patients underwent colorectal SILS procedures. In 74 patients (31.6%), an ileostomy (n = 41) or a colostomy (n = 33) was fashioned. Of these, 20 (27% of all ostomies) were umbilical stomas. The 20 u-stoma patients, 10 men and 10 women, received either a loop ileostomy (n = 14) or an end (n = 4) or loop (n = 2) colostomy. The mean age was 52 years (range 29-81 years). There was no mortality. Operative stoma-related morbidity occurred in only 5% of patients (n = 1: ileal torsion volvulus). Median follow-up after stoma formation was 30 months (range 12-59 months). Adjustment to the stoma and quality of life were satisfactory as estimated by both the patient and the stoma therapist. All stomas were reversed. At a median follow-up of 27.5 months (range 7-55 months) after stoma reversal, two patients had reoperation for incisional hernia. CONCLUSION: This preliminary experience showed that u-stoma is a feasible and safe alternative to more conventional ostomy after SILS.

Effect of water restriction on milk yield and quality in Lacaune breed ewes.

The effect of water restriction on body weight, body condition score, milk yield, and milk composition and rheological characteristics in intensively reared Lacaune ewes was evaluated. After 7 days of adaptation, the trial lasted 28 days. Thirty lactating ewes (48 ± 5 months of age; mean value ± standard deviation) at the beginning of third lactation month were divided into three groups (n = 10), corresponding to the following water restriction treatment: a group control received no drinking water restriction (W100), and two groups received water to the extent of 80% and 60% of W100 daily consumption (W80 and W60 group respectively). The effects of water restriction were assessed at the beginning of experiment (D0), at D14 and D28. The W60 group resulted in a significant decrease in body weight, body condition score, milk yield and feed consumption of hay due to the experimental treatment; whereas a marked increase in both W60 and W80 groups of milk lactose, urea, sodium, sodium chloride content and titratable acidity was observed. Rheological parameters of milk, rennet coagulation time and curd-firming time were positively affected by water restriction treatments, with a decrease in both experimental groups of the time required for the formation of a stable and firm curd. Results highlight the importance of water consumption in dairy sheep. The scarce availability of water, significantly affecting ewes milk production, is worthy of particular attention in arid area where water stress-resistant breeds should be considered. This study underlines that milk yield, being closely linked to the availability of water of the breeding habitat due to its high water content (about 81%), could be reached in area where water is not a limiting factor without reducing the genetic expression of the animals. Less severe water restrictions, such as 20% of daily voluntary water intake, produce no detrimental effect on milk yield.

Does DCD Donor Time-to-Death Affect Recipient Outcomes? Implications of Time-to-Death at a High-Volume Center in the United States.

For donation after circulatory death (DCD), many centers allow 1 h after treatment withdrawal to donor death for kidneys. Our center has consistently allowed 2 h. We hypothesized that waiting longer would be associated with worse outcome. A single-center, retrospective analysis of DCD kidneys transplanted between 2008 and 2013 as well as a nationwide survey of organ procurement organization DCD practices were conducted. We identified 296 DCD kidneys, of which 247 (83.4%) were transplanted and 49 (16.6%) were discarded. Of the 247 recipients, 225 (group 1; 91.1%) received kidneys with a time to death (TTD) of 0-1 h; 22 (group 2; 8.9%) received grafts with a TTD of 1-2 h. Five-year patient survival was 88.8% for group 1, and 83.9% for group 2 (p = 0.667); Graft survival was also similar, with 5-year survival of 74.1% for group 1, and 83.9% for group 2 (p = 0.507). The delayed graft function rate was the same in both groups (50.2% vs. 50.0%, p = 0.984). TTD was not predictive of graft failure. Nationally, the average maximum wait-time for DCD kidneys was 77.2 min. By waiting 2 h for DCD kidneys, we performed 9.8% more transplants without worse outcomes. Nationally, this practice would allow for hundreds of additional kidney transplants, annually.

Choosing the individual rehabilitation program for patients with intermittent claudication.

This study is aimed at identifying the collateral circulation in case of femoral-aorta-iliac axis obstruction, with the purpose of a more correct therapeutic indication being either medical or surgical or physiotherapeutic or combined.

Metabolomics of AS-5 RBC supernatants following routine storage.

BACKGROUND AND OBJECTIVES: The safety and efficacy of stored red blood cells (RBCs) transfusion has been long debated due to retrospective clinical evidence and laboratory results, indicating a potential correlation between increased morbidity and mortality following transfusion of RBC units stored longer than 14 days. We hypothesize that storage in Optisol additive solution-5 leads to a unique metabolomics profile in the supernatant of stored RBCs. MATERIALS AND METHODS: Whole blood was drawn from five healthy donors, RBC units were manufactured, and prestorage leucoreduced by filtration. Samples were taken on days 1 and 42, the cells removed, and mass spectrometry-based metabolomics was performed. RESULTS: The results confirmed the progressive impairment of RBC energy metabolism by day 42 with indirect markers of a parallel alteration of glutathione and NADPH homeostasis. Moreover, oxidized pro-inflammatory lipids accumulated by the end of storage. CONCLUSION: The supernatants from stored RBCs may represent a burden to the transfused recipients from a metabolomics standpoint.

Enteric glia: A new player in inflammatory bowel diseases.

In addition to the well-known involvement of macrophages and neutrophils, other cell types have been recently reported to substantially contribute to the onset and progression of inflammatory bowel diseases (IBD). Enteric glial cells (EGC) are the equivalent cell type of astrocyte in the central nervous system (CNS) and share with them many neurotrophic and neuro-immunomodulatory properties. This short review highlights the role of EGC in IBD, describing the role played by these cells in the maintenance of gut homeostasis, and their modulation of enteric neuronal activities. In pathological conditions, EGC have been reported to trigger and support bowel inflammation through the specific over-secretion of S100B protein, a pivotal neurotrophic factor able to induce chronic inflammatory changes in gut mucosa. New pharmacological tools that may improve the current therapeutic strategies for inflammatory bowel diseases (IBD), lowering side effects (i.e. corticosteroids) and costs (i.e. anti-TNFα monoclonal antibodies) represent a very important challenge for gastroenterologists and pharmacologists. Novel drugs capable to modulate enteric glia reactivity, limiting the pro-inflammatory release of S100B, may thus represent a significant innovation in the field of pharmacological interventions for inflammatory bowel diseases.

Comparative characteristics of DNA polymorphisms of κ-casein gene (CSN3) in the horse and donkey.

The aims of this study were to assess the genetic variability in the exon 1 of the κ-casein gene in four Italian horse populations (Italian Saddle horse, Italian Trotter, Italian Heavy Draught horse, and Murgese horse) and in a sample of Martina Franca donkey by estimating genotype, allele and haplotype frequencies, as well as several population genetic indices. Genotyping of the selected polymorphisms was performed using the PCR-RFLP technique with two restriction enzymes: PstI and BseYI aimed to discover the presence of c.-66A>G and c.-36C>A polymorphism, respectively. Both these loci were found to be polymorphic in horses with some differences depending on the breed. No genetic variability was observed in Martina Franca donkey breed. In the equine species no selective pressure for milk purpose was performed, therefore the polymorphisms at milk protein loci were mainly considered as result of natural selection or as indirect consequence of selection oriented to increase body size or to improve conformation. From this point of view these two single nucleotide polymorphisms and particularly the c.-36C>A one could be useful instruments for population studies.

Short communication: determination of lactoferrin in Feta cheese whey with reversed-phase high-performance liquid chromatography.

In the current paper, a method is introduced to determine lactoferrin in sweet whey using reversed-phase HPLC without any pretreatment of the samples or use of a separation technique. As a starting point, the most common HPLC protocols for acid whey, which included pretreatment of the whey along with a sodium dodecyl sulfate-PAGE step, were tested. By skipping the pretreatment and the separation steps while altering the gradient profile, different chromatographs were obtained that proved to be equally efficient to determine lactoferrin. For this novel 1-step reversed-phase HPLC method, repeatability was very high over a wide range of concentrations (1.88% intraday to 5.89% interday). The limit of detection was 35.46µg/mL [signal:noise ratio (S/N)=3], whereas the limit of quantification was 50.86µg/mL (S/N=10). Omitting the pretreatment step caused a degradation of the column's lifetime (to approximately 2,000 samples). As a result, the lactoferrin elution time changed, but neither the accuracy nor the separation ability of the method was significantly influenced. We observed that this degradation could be easily avoided or detained by centrifuging the samples to remove fat or by extensive cleaning of the column after every 5 samples.

Complete absence of GLUT1 does not impair human terminal erythroid differentiation.

The Glucose transporter 1 (GLUT1) is one of the most abundant proteins within the erythrocyte membrane and is required for glucose and dehydroascorbic acid (Vitamin C precursor) transport. It is widely recognized as a key protein for red cell structure, function, and metabolism. Previous reports highlighted the importance of GLUT1 activity within these uniquely glycolysis-dependent cells, in particular for increasing antioxidant capacity needed to avoid irreversible damage from oxidative stress in humans. However, studies of glucose transporter roles in erythroid cells are complicated by species-specific differences between humans and mice. Here, using CRISPR-mediated gene editing of immortalized erythroblasts and adult CD34+ hematopoietic progenitor cells, we generate committed human erythroid cells completely deficient in expression of GLUT1. We show that absence of GLUT1 does not impede human erythroblast proliferation, differentiation, or enucleation. This work demonstrates for the first-time generation of enucleated human reticulocytes lacking GLUT1. The GLUT1-deficient reticulocytes possess no tangible alterations to membrane composition or deformability in reticulocytes. Metabolomic analyses of GLUT1-deficient reticulocytes reveal hallmarks of reduced glucose import, downregulated metabolic processes and upregulated AMPK-signalling, alongside alterations in antioxidant metabolism, resulting in increased osmotic fragility and metabolic shifts indicative of higher oxidant stress. Despite detectable metabolic changes in GLUT1 deficient reticulocytes, the absence of developmental phenotype, detectable proteomic compensation or impaired deformability comprehensively alters our understanding of the role of GLUT1 in red blood cell structure, function and metabolism. It also provides cell biological evidence supporting clinical consensus that reduced GLUT1 expression does not cause anaemia in GLUT1 deficiency syndrome.

Mitochondrial reverse electron transport in myeloid cells perpetuates neuroinflammation.

Sustained smouldering, or low grade, activation of myeloid cells is a common hallmark of several chronic neurological diseases, including multiple sclerosis (MS) 1 . Distinct metabolic and mitochondrial features guide the activation and the diverse functional states of myeloid cells 2 . However, how these metabolic features act to perpetuate neuroinflammation is currently unknown. Using a multiomics approach, we identified a new molecular signature that perpetuates the activation of myeloid cells through mitochondrial complex II (CII) and I (CI) activity driving reverse electron transport (RET) and the production of reactive oxygen species (ROS). Blocking RET in pro-inflammatory myeloid cells protected the central nervous system (CNS) against neurotoxic damage and improved functional outcomes in animal disease models in vivo . Our data show that RET in myeloid cells is a potential new therapeutic target to foster neuroprotection in smouldering inflammatory CNS disorders 3 .

Haemoglobin glycation (Hb1Ac) increases during red blood cell storage: a MALDI-TOF mass-spectrometry-based investigation.

Haemoglobin A1c (HbA1c ) represents a key biomarker in diabetes diagnosis and management, as it is indicative of recent blood glucose concentrations. Glycation of haemoglobin is a non-enzymatic irreversible process that is promoted by the prolonged exposure of erythrocytes to high glucose concentrations, a condition that is known to occur under blood banking conditions. However, controversial data indicate no clear hint as to whether and to which extent HbA1c accumulates during red blood cell storage. Hereby, we propose the application of a validated MALDI-TOF mass-spectrometry-based method to this issue and report the observation about HbA1c levels apparently increasing over storage progression.

Short-term outcomes after rehabilitation treatment in patients selected by a novel rehabilitation score system (Brusciano score) with or without previous stapled transanal rectal resection (STARR) for rectal outlet obstruction.

PURPOSE: The purpose of this study was to examine short-term outcomes of rehabilitation treatment in patients with or without previous stapled transanal resection (STARR) for rectal outlet obstruction by using a novel rehabilitation score system (Brusciano score). METHODS: This is a retrospective cohort study conducted at a single tertiary referral institution including all patients with chronic functional constipation admitted to the outpatient unit from 2004 to 2009. RESULTS: Among 330 consecutive patients, 247 (74.8 %) (204 females and 43 males) showing a significantly higher rehabilitation score (mean of 15.7 ± 1.8; range, 7-25) than healthy controls (mean, 3.2 ± 1.2; range 2-6) (p < .0001) were selected for rehabilitation. Of the 247 patients evaluated, group A (no previous surgery) consisted of 170 patients (53 males; mean age, 44.8 ± 12.9 years; range, 19-80) of which 38 presented mixed constipation, whereas group B (previous surgery) consisted of 77 patients (18 males; mean age, 47.0 ± 11.2 years; range, 22-81). The Brusciano score, Agachan-Wexner score and quality of life improved in both groups of patients after treatment. Better improvements of Brusciano and Agachan-Wexner scores were observed in patients with previous STARR (group B). CONCLUSIONS: The rehabilitation score system employed in this study seems to be a useful tool in selecting and assessing the outcome of patients who might benefit from rehabilitation treatment. Constipation and quality of life were significantly improved by the rehabilitation treatment. Further studies are needed to clarify either the impact of rehabilitation treatment on long-term outcome of patients treated for rectal outlet obstruction or its role in those who develop problems over time.

Improvements in one severe progressive multiple sclerosis patient quality of life after an intensity fluid dynamic treatment.

Clinical case: A 49-year-old man (MM72) affected by Secondary Progressive Multiple Sclerosis (SP-MS) since 1998. On last 3 years, neurologists valued 9.0 the patient MM72's EDSS. Methods: MM72 was treated by acoustic waves, modulated in frequency and power by the MAM device, according to an ambulatory intensive protocol. Patient's treatments schedule was organized in thirty cycles of DrenoMAM and AcuMAM, and manual cervical spinal adjustments. Before and after treatments, MSIS-29, Barthel, FIM, EDSS, ESS, and FSS questionnaires were administered to the patient. Results: MM72 patient had improvements in all index score (MSIS-29, Barthel, FIM, EDSS, ESS and FSS) after 30 treatments by MAM plus cervical spine chiropractic adjustments. He showed a significative improvement of his disability and the restore of many functions. After MAM treatments, MM72's cognitive sphere improved of 370%. Fur-thermore, after 5 years of paraplegy, he regained his lower limbs and feet fingers movements with an increase of 230%. Conclusion: We suggest ambulatory intensive treatments by fluid dynamic MAM protocol in SP-MS patients. Statistical analyses are in progress on a larger sample of SP-MS patients.

The impact of social, cognitive and attitudinal dimensions on college students' support for organ donation.

This study investigates how college students can be social support catalysts for organ donation and how social, cognitive and attitudinal dimensions impact organ donor registration. A total of 317 people participated in the exploratory portion of the project and a total of 1800 responses were obtained from an online survey to members of a national student organization. The findings show that perceptions of the benefits of organ donation and altruistic motives had the greatest impact on the support for organ donation while respondents' knowledge about how to register to be an organ donor was the dominant dimension for donor registration status. Social-based communications had the next greatest impact for both support and donor registration. Based on the findings, an 18-month social media campaign was launched with the student organization that had 20 421 website visitors, 4473 Facebook members, 1189 YouTube video submissions with 164 000 views, motivated 19 623 people to go to a state's organ donor registration page, and had 9000 documented organ donor registrations. Within the student organization, organ donor registration increased by 28%. On the basis of these project results, Donate Life America and other sponsors have provided funding for two additional years.

Red blood cell storage and cell morphology.

AIM: In this study, we performed weekly assessment of morphology-related parameters through monitoring of CPD-SAGM leuco-filtered erythrocyte concentrates from blood withdrawal until the 42nd day of storage. BACKGROUND: Liquid storage of red blood cells (RBCs) delivers a blood-derived therapeutic, which is safe, available, effective and affordable for most patients who need transfusion therapy in developed countries. However, a growing body of accumulating controversial evidences, from either biochemical or retrospective clinical studies, prompted safety concerns about longer stored RBCs. METHODS: Statistical image analysis through scanning electron microscope was coupled to osmotic fragility and erythrocyte sedimentation rate. RESULTS: We could observe that by day 21 more than 50% of RBCs displayed non-discocyte phenotypes. This observation was related to an increase in osmotic fragility, which was totally overlapped in day 0 controls and day 7 RBCs while only slightly augmented in day 14 samples. Cation dysregulation (pH internal/external alteration and potassium) might both reflect and trigger a negative feedback loop with metabolic fluxes and membrane cation pumps. CONCLUSION: Morphology parameters suggest that significant alterations to RBC morphology over storage duration occur soon after the 14th day of storage, as to become significant enough within the 21st day.

Role of pre and post-operative oral calcium and vitamin D supplements in prevention of hypocalcemia after total thyroidectomy.

AIM: to evaluate the role of pre and post-operative oral calcium and vitamin D supplements in prevention of hypocalcemia after total thyroidectomy. PATIENTS AND METHODS: 50 consecutive patients, undergoing total thyroidectomy, were enrolled. Oral calcium and vitamin D were administered in the pre and post-operative time. The data concerning symptomatic and laboratoristic hypocalcemia were collected. RESULTS: Incidence of symptomatic hypocalcemia was very low (6%); incidence of laboratoristic hypocalcemia was 10%. No permanent hypocalcemia developed. CONCLUSIONS: Implementing oral calcium and vitamin D both before and after total thyroidectomy can reduce the incidence of hypocalcemia related to surgery.