Pneumocystis murina lesions in lungs of experimentally infected Cd40l-/- mice.

The Cd40l-/- mouse is a well-established model of X-linked hyper-immunoglobulin M (IgM) syndrome, an immunodeficiency disorder of human beings characterized by the lack of expression of the CD40 ligand (CD40L) on activated T-cells, predisposing to infections with opportunistic pathogens like Pneumocystis jirovecii. The aim of our study was to describe the pulmonary lesions in Cd40l-/- mice experimentally infected with Pneumocystis murina, in comparison with naturally infected severe combined immunodeficient (SCID) mice. Formalin-fixed paraffin-embedded lungs from 26 Cd40l-/-, 11 SCID, and 5 uninfected Cd40l-/- mice were examined by histology and immunohistochemistry for the presence of the pathogen and for leukocyte populations (CD3, CD4, CD45R/B220, CD8a, Iba-1, Ly-6G, CD206, MHC II, and NKp46/NCR1). Infection was confirmed by immunohistochemistry in 18/26 (69%) Cd40l-/- mice and in 11/11 (100%) SCID mice. Fourteen out of 26 (54%) Cd40l-/- mice had interstitial pneumonia. Twenty-three out of 26 (88%) Cd40l-/- mice had peribronchiolar/perivascular lymphoplasmacytic infiltrates, rich in B-cells and Mott cells. Acidophilic macrophage pneumonia was additionally found in 20/26 (77%) Cd40l-/- mice. Only 4/11 (36%) SCID mice had interstitial pneumonia, but no peribronchiolar/perivascular infiltrates or acidophilic macrophage pneumonia were observed in this strain. This study represents the first description of pulmonary histopathological lesions in Cd40l-/- mice infected with P. murina. We speculate that the singular characteristics of the inflammatory infiltrates observed in Cd40l-/- mice could be explained by the specific immune phenotype of the model.

Caloric restriction and metformin selectively improved LKB1-mutated NSCLC tumor response to chemo- and chemo-immunotherapy.

BACKGROUND: About 10% of NSCLCs are mutated in KRAS and impaired in STK11/LKB1, a genetic background associated with poor prognosis, caused by an increase in metastatic burden and resistance to standard therapy. LKB1 is a protein involved in a number of biological processes and is particularly important for its role in the regulation of cell metabolism. LKB1 alterations lead to protein loss that causes mitochondria and metabolic dysfunction that makes cells unable to respond to metabolic stress. Different studies have shown how it is possible to interfere with cancer metabolism using metformin and caloric restriction (CR) and both modify the tumor microenvironment (TME), stimulating the switch from "cold" to "hot". Given the poor therapeutic response of KRASmut/LKB1mut patients, and the role of LKB1 in cell metabolism, we examined whether the addition of metformin and CR enhanced the response to chemo or chemo-immunotherapy in LKB1 impaired tumors. METHODS: Mouse cell lines were derived from lung nodules of transgenic mice carrying KRASG12D with either functional LKB1 (KRASG12D/LKB1wt) or mutated LKB1 (KRASG12D/LKB1mut). Once stabilized in vitro, these cell lines were inoculated subcutaneously and intramuscularly into immunocompetent mice. Additionally, a patient-derived xenograft (PDX) model was established by directly implanting tumor fragments from patient into immunocompromised mice. The mice bearing these tumor models were subjected to treatment with chemotherapy or chemo-immunotherapy, both as standalone regimens and in combination with metformin and CR. RESULTS: Our preclinical results indicate that in NSCLC KRASmut/LKB1mut tumors, metformin and CR do enhance the response to chemo and chemo-immunotherapy, inducing a metabolic stress condition that these tumors are not able to overcome. Analysis of immune infiltrating cells did not bring to light any strong correlation between the TME immune-modulation and the tumor response to metformin and CR. CONCLUSION: Our in vitro and in vivo preliminary studies confirm our hypothesis that the addition of metformin and CR is able to improve the antitumor activity of chemo and chemoimmunotherapy in LKB1 impaired tumors, exploiting their inability to overcome metabolic stress.

Chemotherapy-induced neutropenia elicits metastasis formation in mice by promoting proliferation of disseminated tumor cells.

Chemotherapy is the standard of care for most malignancies. Its tumor debulking effect in adjuvant or neoadjuvant settings is unquestionable, although secondary effects have been reported that paradoxically promote metastasis. Chemotherapy affects the hematopoietic precursors leading to myelosuppression, with neutropenia being the main hematological toxicity induced by cytotoxic therapy. We used renal and lung murine tumor models metastatic to the lung to study chemotherapy-induced neutropenia (CIN) in the metastatic process. Cyclophosphamide and doxorubicin, two myelosuppressive drugs, but not cisplatin, increased the burden of artificial metastases to the lung, by reducing neutrophils. This effect was recapitulated by treatment with anti-Ly6G, the selective antibody-mediated neutrophil depletion that unleashed the formation of lung metastases in both artificial and spontaneous metastasis settings. The increased cancer dissemination was reversed by granulocyte-colony stimulating factor-mediated boosting of neutrophils in combination with chemotherapy. CIN affected the early metastatic colonization of the lung, quite likely promoting the proliferation of tumor cells extravasated into the lung at 24-72 hours. CIN did not affect the late events of the metastatic process, with established metastasis to the lung, nor was there any effect on the release of cancer cells from the primary, whose growth was, in fact, somewhat inhibited. This work suggests a role of neutrophils associated to a common cancer treatment side effect and claims a deep dive into the relationship between chemotherapy-induced neutropenia and metastasis.

Chronic cholesterol administration to the brain supports complete and long-lasting cognitive and motor amelioration in Huntington's disease.

Evidence that Huntington's disease (HD) is characterized by impaired cholesterol biosynthesis in the brain has led to strategies to increase its level in the brain of the rapidly progressing R6/2 mouse model, with a positive therapeutic outcome. Here we tested the long-term efficacy of chronic administration of cholesterol to the brain of the slowly progressing zQ175DN knock-in HD mice in preventing ("early treatment") or reversing ("late treatment") HD symptoms. To do this we used the most advanced formulation of cholesterol loaded brain-permeable nanoparticles (NPs), termed hybrid-g7-NPs-chol, which were injected intraperitoneally. We show that one cycle of treatment with hybrid-g7-NPs-chol, administered in the presymptomatic ("early treatment") or symptomatic ("late treatment") stages is sufficient to normalize cognitive defects up to 5 months, as well as to improve other behavioral and neuropathological parameters. A multiple cycle treatment combining both early and late treatments ("2 cycle treatment") lasting 6 months generates therapeutic effects for more than 11 months, without severe adverse reactions. Sustained cholesterol delivery to the brain of zQ175DN mice also reduces mutant Huntingtin aggregates in both the striatum and cortex and completely normalizes synaptic communication in the striatal medium spiny neurons compared to saline-treated HD mice. Furthermore, through a meta-analysis of published and current data, we demonstrated the power of hybrid-g7-NPs-chol and other strategies able to increase brain cholesterol biosynthesis, to reverse cognitive decline and counteract the formation of mutant Huntingtin aggregates. These results demonstrate that cholesterol delivery via brain-permeable NPs is a therapeutic option to sustainably reverse HD-related behavioral decline and neuropathological signs over time, highlighting the therapeutic potential of cholesterol-based strategies in HD patients. DATA AVAILABILITY: This study does not include data deposited in public repositories. Data are available on request to the corresponding authors.

Re-evaluation of the risks to public health related to the presence of bisphenol A (BPA) in foodstuffs.

In 2015, EFSA established a temporary tolerable daily intake (t-TDI) for BPA of 4 µg/kg body weight (bw) per day. In 2016, the European Commission mandated EFSA to re-evaluate the risks to public health from the presence of BPA in foodstuffs and to establish a tolerable daily intake (TDI). For this re-evaluation, a pre-established protocol was used that had undergone public consultation. The CEP Panel concluded that it is Unlikely to Very Unlikely that BPA presents a genotoxic hazard through a direct mechanism. Taking into consideration the evidence from animal data and support from human observational studies, the immune system was identified as most sensitive to BPA exposure. An effect on Th17 cells in mice was identified as the critical effect; these cells are pivotal in cellular immune mechanisms and involved in the development of inflammatory conditions, including autoimmunity and lung inflammation. A reference point (RP) of 8.2 ng/kg bw per day, expressed as human equivalent dose, was identified for the critical effect. Uncertainty analysis assessed a probability of 57-73% that the lowest estimated Benchmark Dose (BMD) for other health effects was below the RP based on Th17 cells. In view of this, the CEP Panel judged that an additional uncertainty factor (UF) of 2 was needed for establishing the TDI. Applying an overall UF of 50 to the RP, a TDI of 0.2 ng BPA/kg bw per day was established. Comparison of this TDI with the dietary exposure estimates from the 2015 EFSA opinion showed that both the mean and the 95th percentile dietary exposures in all age groups exceeded the TDI by two to three orders of magnitude. Even considering the uncertainty in the exposure assessment, the exceedance being so large, the CEP Panel concluded that there is a health concern from dietary BPA exposure.

Tobacco Seed-Based Oral Vaccination against Verocytotoxic O138 Escherichia coli as Alternative Approach to Antibiotics in Weaned Piglets.

Post-weaning diarrhoea and enterotoxaemia caused by Escherichia coli are serious threats in the pig (Sus scrofa domesticus) livestock industry and are responsible for economic losses related to mortality, morbidity and stunted growth. The aim of this study was to evaluate the effect of an engineered tobacco seeds-based edible vaccine in O138 Escherichia coli-challenged piglets throughout a multidisciplinary approach. Thirty-six weaned piglets were enrolled and randomly divided into two experimental groups, a control (C; n = 18) group and a tobacco edible vaccination group (T, n = 18), for 29 days of trial. At days 0, 1, 2, 5 and 14, piglets of the T group were fed with 10 g of the engineered tobacco seeds line expressing F18 and VT2eB antigens, while the C group received wild-type tobacco seeds. After 20 days, 6 piglets/group were orally challenged with the Escherichia coli O138 strain (creating four subgroups: UC = unchallenged control, CC = challenged control, UT = unchallenged tobacco, CT = challenged tobacco) and fed with a high protein diet for 3 consecutive days. Zootechnical, clinical, microbiological, histological and immunological parameters were assayed and registered during the 9 days of post-challenge follow up. At 29 days post-challenge, the CT group displayed a lower average of the sum of clinical scores compared to the CC group (p < 0.05), while the CC group showed a higher average sum of the faecal score (diarrhoea) (p < 0.05) than the CT group. A decreased number of days of shedding of the pathogenic strain was observed in the CT compared to the CC group (p < 0.05). Specific anti-F18 IgA molecules were significantly higher in the CT group compared to the CC group's faecal samples during the post-challenge period (p < 0.01). In conclusion, edible vaccination with engineered tobacco seeds showed a protective effect on clinical symptoms and diarrhoea incidence during the post-challenge period, characterized by a limited time of pathogenic strain shedding in faeces.

Retrospective study of tumors from cattle slaughtered in Lombardy (Italy): preliminary evaluation on the establishment of a bovine cancer registry.

Forty­one tumors were detected in a population of 1,649,003 cattle slaughtered in 4 abattoirs in Lombardy over a 5­year period, for an overall prevalence of 2.5 tumors per 100,000 cattle. Tumors were classified according to the WHO histological classification of tumors of domestic animals. Alimentary and hemopoietic systems were commonly affected with 9 cases each. Other affected sites were the respiratory (n = 3), urinary (n = 2), endocrine (n = 2), musculoskeletal (n = 2), nervous (n = 1), and cardiovascular (n = 1) systems. The peritoneum was affected by 6 cases, while the primary location of 3 tumors of the connective tissues and 3 metastatic carcinomas was unidentified. Liver tumors and mesotheliomas, for which environmental risk factors are well­known in humans, were common, as well as tumors typically encountered in pediatric human patients (tumors of mesenchymal tissues, pulmonary blastomas and nephroblastomas). These findings suggest the useful role of bovines as sentinel and model for human carcinogenesis. Our study indicates that the establishment of a bovine cancer registry in Lombardy is feasible considering its potential contribution to understanding the role of environmental risk factors in the genesis of tumors in animals and humans.

Cranial Spinal Spreading of Canine Brain Gliomas after Hypofractionated Volumetric-Modulated Arc Radiotherapy and Concomitant Temozolomide Chemotherapy: A Four-Case Report.

Gliomas are the second-most-common primary brain tumors in dogs. Surgery and radiotherapy are established treatment approaches with similar median survival time, whereas conventional chemotherapy is burdened by severe adverse effects. Spinal and leptomeningeal spread of gliomas have been described following radiotherapy treatment alone. The purpose of this study was to evaluate the outcome for four dogs with primary high-grade gliomas in the forebrain without evidence, at diagnosis, of neoplastic invasion along the spinal cord, that were treated with concomitant chemotherapy (temozolomide) and hypofractionated volumetric-modulated arc radiotherapy (VMAT-RT). Temozolomide was selected for its radiosensitive properties, and radiotherapy dose protocols of 37 Gy in 7 fractions or 42 Gy in 10 fractions were used. After an initial complete or partial response, tumors recurred across the cranial-spinal pathway. Post-mortem macroscopic examinations confirmed swollen spinal cord and hyperemic meningeal sleeve, with nodular lesions on the meningeal surface. Microscopically, infiltration of the spinal cord and meninges by neoplastic cells (with features of oligodendrogliomas) were observed. This work seems to suggest that the entire central nervous system should be investigated in diagnostic examinations of canine gliomas. Dose-escalation trials and/or spinal cord prophylaxis treatment could also be evaluated to prevent tumor progression.

EMILIN-1 deficiency promotes chronic inflammatory disease through TGFß signaling alteration and impairment of the gC1q/α4ß1 integrin interaction.

Alterations in extracellular matrix (ECM) components that modulate inflammatory cell behavior have been shown to serve as early starters for multifactorial diseases such as fibrosis and cancer. Here, we demonstrated that loss of the ECM glycoprotein EMILIN-1 alters the inflammatory context in skin during IMQ-induced psoriasis, a disease characterized by a prominent inflammatory infiltrate and alteration of vessels that appear dilated and tortuous. Abrogation of EMILIN-1 expression or expression of the EMILIN-1 mutant E933A impairs macrophage polarization and leads to imbalanced tissue homeostasis. We found that EMILIN-1 deficiency is associated with dilated lymphatic vessels, increased macrophage recruitment and psoriasis severity. Importantly, the null or mutant EMILIN-1 background was characterized by the induction of a myofibroblast phenotype, which in turn drove macrophages towards the M1 phenotype. By using the transgenic mouse model carrying the E933A mutation in the gC1q domain of EMILIN-1, which abolishes the interaction with α4- and α9-integrins, we demonstrated that the observed changes in TGFß signaling were due to both the EMI and gC1q domains of EMILIN-1. gC1q may exert multiple functions in psoriasis, in the context of a final, more consistent inflammatory condition by controlling skin homeostasis via interaction with both keratinocytes and fibroblasts, influencing non-canonical TGFß signaling, and likely acting on lymphatic vessel structure and function. The analyses of human psoriatic lesions, in which lower levels of EMILIN-1 were present with a very rare association with lymphatic vessels, support the multifaceted role of this ECM component in the skin inflammatory scenario.

IL-1R8 Downregulation and Concomitant TLR7 and TLR9 Upregulation Are Related to the Pathogenesis of Canine Diffuse Large B-Cell Lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is the most common hematological malignancy in humans and dogs. Several studies disclosed some similarities between the two species, including the constitutive activation of the NF-κB pathway as a fundamental underlying pathogenetic mechanism. In humans, the downregulation of IL-1R8 is implicated in DLBCL development, but its role in dogs has not been explored so far. To gain insight into the pathogenesis of this tumor in dogs, we evaluated the mRNA and protein expression of IL-1R8 in 12 control lymph nodes obtained from dogs not bearing tumors and from 50 dogs with DLBCL. Moreover, we analyzed through qRT-PCR the expression of TLR7, TLR9, MYC, and p52 genes that are known to be involved in the IL-1R8 regulatory network. IL-1R8 and p52 were downregulated in DLBCLs compared to control lymph nodes (p < 0.001), while a higher expression of TLR7, TLR9, and MYC was observed in DLBCLs (p < 0.01). Immunohistochemistry confirmed the gene expression results, revealing a significantly lower IL-1R8 staining score in DLBCLs compared to control lymph nodes (p < 0.0001). Taken together, these results suggest that IL-1R8 downregulation may represent one of the mechanisms driving DLBCL pathogenesis in dogs, mainly through the dysregulation of the Toll-like/interleukin receptors signaling cascade and the aberrant activation of the classical NF-κB pathway.

Lack of ApoA-I in ApoEKO Mice Causes Skin Xanthomas, Worsening of Inflammation, and Increased Coronary Atherosclerosis in the Absence of Hyperlipidemia.

BACKGROUND: HDL (high-density lipoprotein) and its major protein component, apoA-I (apolipoprotein A-I), play a unique role in cholesterol homeostasis and immunity. ApoA-I deficiency in hyperlipidemic, atheroprone mice was shown to drive cholesterol accumulation and inflammatory cell activation/proliferation. The present study was aimed at investigating the impact of apoA-I deficiency on lipid deposition and local/systemic inflammation in normolipidemic conditions. METHODS: ApoE deficient mice, apoE/apoA-I double deficient (DKO) mice, DKO mice overexpressing human apoA-I, and C57Bl/6J control mice were fed normal laboratory diet until 30 weeks of age. Plasma lipids were quantified, atherosclerosis development at the aortic sinus and coronary arteries was measured, skin ultrastructure was evaluated by electron microscopy. Blood and lymphoid organs were characterized through histological, immunocytofluorimetric, and whole transcriptome analyses. RESULTS: DKO were characterized by almost complete HDL deficiency and by plasma total cholesterol levels comparable to control mice. Only DKO showed xanthoma formation and severe inflammation in the skin-draining lymph nodes, whose transcriptome analysis revealed a dramatic impairment in energy metabolism and fatty acid oxidation pathways. An increased presence of CD4+ T effector memory cells was detected in blood, spleen, and skin-draining lymph nodes of DKO. A worsening of atherosclerosis at the aortic sinus and coronary arteries was also observed in DKO versus apoE deficient. Human apoA-I overexpression in the DKO background was able to rescue the skin phenotype and halt atherosclerosis development. CONCLUSIONS: HDL deficiency, in the absence of hyperlipidemia, is associated with severe alterations of skin morphology, aortic and coronary atherosclerosis, local and systemic inflammation.

Bacteriological and pathological investigations on the preputial glands of one-year-old C57BL/6NCrl mice maintained in individually ventilated cages.

Spontaneous infections of the preputial glands represent overlooked health problems in mice that could raise welfare concerns and potentially confound scientific experiments. Agents involved in preputial gland infections have rarely been investigated, with opportunistic pathogens of laboratory animals usually detected in inflamed preputial glands. The aim of this study was to investigate the prevalence of bacterial infection in the preputial glands and the relationship between haematological and pathological changes and infection status. We analysed 40 preputial glands from 20 one-year-old C57BL/6NCrl male mice by using bacteriology, haematology and pathology. Bacteria were isolated from 16/20 (80%) mice, for a total of 32/40 (80%) examined preputial glands. Enterobacter cloacae, Pasteurella spp., Klebsiella spp. and Staphylococcus aureus were identified in 35%, 17.5%, 15% and 12.5% of the examined glands, respectively. Preputial gland inflammation was identified in 29/40 (72.5%) glands and was classified as chronic interstitial adenitis in 27 cases and suppurative adenitis in the remaining two glands. No haematological changes were found in mice with infected glands. Histologically, the presence of intralesional bacteria, intraluminal necrotic material, intraluminal keratin accumulation, interstitial inflammatory cell infiltrate and granulocytes (intraluminal and/or interstitial), along with total inflammatory score and total histopathological score, were significantly increased in infected glands and correlated with the bacterial load. Most severe inflammatory changes were identified after S. aureus infection, while ductal hyperkeratosis was significantly increased in glands infected with Klebsiella spp. In conclusion, preputial gland infection was a common event in one-year-old C57BL/6NCrl mice, and bacterial load correlated with pathological findings, while systemic effects were not highlighted by haematology.

Senescent Thyrocytes, Similarly to Thyroid Tumor Cells, Elicit M2-like Macrophage Polarization In Vivo.

Inflammation plays a critical role in thyroid cancer onset and progression. We previously characterized the in vitro interplay between macrophages and senescent human thyrocytes and thyroid tumor-derived cell lines, modeling the early and the late thyroid tumor phases, respectively. We reported that both models are able to induce pro-tumoral M2-like macrophage polarization, through the activation of the COX2-PGE2 axis. Here, we investigated the presence of macrophage infiltrating cells in mouse xenografts derived from the above described cells models. We showed that subcutaneous injection in immunodeficient mice of both senescent human thyrocytes and thyroid tumor-derived cell lines elicits macrophage recruitment. Furthermore, considering the type of macrophage infiltrate, we observed a stronger infiltration of Arginase I positive cells (M2-like). Overall, these results demonstrate the in vivo capability of senescent and tumor thyroid cells to recruit and polarize macrophages, suggesting that the promotion of a pro-tumoral microenvironment through tumor associated macrophages may occurs in late as well as in early thyroid tumor stages, favoring tumor onset and progression.

Translational Approach to Induce and Evaluate Verocytotoxic E. coli O138 Based Disease in Piglets.

Pig livestock was influenced by several global concerns that imposed a re-thinking of the farming system, which included the reduction in chemical dependency and the development of antimicrobial alternatives. Post-weaning diarrhea and enterotoxaemia caused by Escherichia coli, are serious threats that are responsible for the economic losses related to mortality, morbidity and stunted growth in weaning piglets. The aim of the study was to set up experimental conditions to simulate the simultaneous outbreak of post-weaning diarrhea and enterotoxaemia in weaned piglets, through verocytotoxic O138 Escherichia coli challenge, with a multidisciplinary approach. Eighteen piglets susceptible to F18 VTEC infection were selected by polymerase chain reaction for polymorphism on the fucosyltransferase 1 gene and randomly divided in two experimental groups, non-infected controls (C; n = 6) and infected ones (I; n = 12) and housed into individual pens at the same environmental conditions for 29 days. At day 20, I pigs were orally inoculated with Escherichia coli O138 and fed a high protein ration for 3 days. Zootechnical, clinical, microbiological, histological and immunological parameters were evaluated along the follow up (3 and 9 days). Experimental infection, confirmed by bacteria faecal shedding of the I group, significantly affected the clinical status. The I group showed significantly higher total scores, corresponding to medians of the sum of daily scores from days 1 to 3 (Σ3) and 1 to 9 (Σ9) post infection, epiphora, vitality, hair irregularity, oedema and depression. Histological examination showed evident inflammatory infiltrate of lymphocytes, and follicular hyperplasia in I pigs; in the same group, the immunohistochemical and immunological assays revealed an increase in IgG in the intestinal crypts and CD3-positive T cells in intestinal epithelium. The experimental Escherichia coli infection in controlled conditions is crucial for both the evaluation of innovative compounds and the elucidation of the mechanisms associated with the persistence of antibacterial resistant strains. In conclusion, the adopted infection model, carried out on receptor-mediated susceptible piglets, allowed us to identify a discriminative panel of clinical symptoms related to Escherichia coli O138 infection, and could be used to assess the protective effect of antibiotic alternatives.

Repeated oral administration of low doses of silver in mice: tissue distribution and effects on central nervous system.

BACKGROUND: Widespread use of silver in its different forms raises concerns about potential adverse effects after ingestion, the main exposure route for humans. The aim of this study was to investigate in CD-1 (ICR) male mice the tissue distribution and in vivo effects of 4-week oral exposure to 0.25 and 1 mg Ag/kg bw 10 nm citrate coated silver nanoparticles (AgNPs) and 1 mg Ag/kg bw silver acetate (AgAc) at the end of treatment (EoT) and after 4 weeks of recovery. RESULTS: There were no treatment-related clinical signs and mortality, and no significant effects on body and organ weights at the EoT and after recovery. Treatment-related changes in hematology and clinical chemistry were found after recovery, the most relevant being a dose-dependent lymphopenia and increased triglycerides in AgNP-treated mice, and increased levels of urea in all treated groups, associated with decreased albumin only in AgAc-treated mice. At the EoT the highest silver concentration determined by Triple Quadrupole ICP-MS analysis was found in the brain, followed by testis, liver, and spleen; much lower concentrations were present in the small intestine and kidney. Tissue silver concentrations were slightly higher after exposure to AgAc than AgNPs and dose dependent for AgNPs. After recovery silver was still present in the brain and testis, highlighting slow elimination. No histopathological changes and absence of silver staining by autometallography were observed in the organs of treated mice. At the EoT GFAP (astrocytes) immunoreactivity was significantly increased in the hippocampus of AgNP-treated mice in a dose-dependent manner and Iba1 (microglial cells) immunoreactivity was significantly increased in the cortex of 1 mg/kg bw AgNP-treated mice. After recovery, a significant reduction of Iba1 was observed in the cortex of all treated groups. TEM analysis of the hippocampus revealed splitting of basement membrane of the capillaries and swelling of astrocytic perivascular end-feet in 1 mg/kg bw AgNP- and AgAc-treated mice at the EoT. CONCLUSIONS: Our study revealed accumulation and slow clearance of silver in the brain after oral administration of 10 nm AgNPs and AgAc at low doses in mice, associated with effects on glial cells and ultrastructural alterations of the Blood-Brain Barrier.

Esmolol during cardiopulmonary resuscitation reduces neurological injury in a porcine model of cardiac arrest.

Primary vasopressor efficacy of epinephrine during cardiopulmonary resuscitation (CPR) is due to its α-adrenergic effects. However, epinephrine plays ß1-adrenergic actions, which increasing myocardial oxygen consumption may lead to refractory ventricular fibrillation (VF) and poor outcome. Effects of a single dose of esmolol in addition to epinephrine during CPR were investigated in a porcine model of VF with an underlying acute myocardial infarction. VF was ischemically induced in 16 pigs and left untreated for 12 min. During CPR, animals were randomized to receive epinephrine (30 µg/kg) with either esmolol (0.5 mg/kg) or saline (control). Pigs were then observed up to 96 h. Coronary perfusion pressure increased during CPR in the esmolol group compared to control (47 ± 21 vs. 24 ± 10 mmHg at min 5, p < 0.05). In both groups, 7 animals were successfully resuscitated and 4 survived up to 96 h. No significant differences were observed between groups in the total number of defibrillations delivered prior to final resuscitation. Brain histology demonstrated reductions in cortical neuronal degeneration/necrosis (score 0.3 ± 0.5 vs. 1.3 ± 0.5, p < 0.05) and hippocampal microglial activation (6 ± 3 vs. 22 ± 4%, p < 0.01) in the esmolol group compared to control. Lower circulating levels of neuron specific enolase were measured in esmolol animals compared to controls (2[1-3] vs. 21[16-52] ng/mL, p < 0.01). In this preclinical model, ß1-blockade during CPR did not facilitate VF termination but provided neuroprotection.

Total laryngectomy in a cat with a laryngeal peripheral nerve sheath tumor.

OBJECTIVE: To report the surgical technique and outcome of total laryngectomy in a single clinical case. STUDY DESIGN: Case report. ANIMAL: A 5-year-old female spayed domestic shorthair cat. METHODS: A cat presented for acute, severe respiratory distress caused by an invasive laryngeal mass. Incisional biopsy was indicative of sarcoma. Computed tomography of head, neck, and thorax was performed revealing no evidence of metastasis. A total laryngectomy and permanent tracheostomy were performed, and the cat could breathe without difficulties immediately postoperatively. Histopathology confirmed a laryngeal low-grade peripheral nerve sheath tumor (PNST). RESULTS: Surgical margins were free of tumor cells. Surgical revision of the tracheostomy stoma due to obstructive granulation tissue was necessary 24 days after the initial surgery. Nine days after revision surgery, the cat was discharged from the hospital. No evidence of local recurrence or metastasis was detected on repeat computed tomography of the head, neck, and thorax at 6 months, nor on chest radiographs at 12 months postoperatively. At the time of writing (13 months postoperatively), the cat is still alive with a good quality of life. CONCLUSION: Total laryngectomy with permanent tracheostomy allowed the complete removal of an obstructive laryngeal PNST and provided a good quality of life in a cat. CLINICAL SIGNIFICANCE: To the authors' knowledge, this case report represents the first detailed description of the surgical procedure and clinical outcome for a total laryngectomy in a cat.

Modeling, optimization, and comparable efficacy of T cell and hematopoietic stem cell gene editing for treating hyper-IgM syndrome.

Precise correction of the CD40LG gene in T cells and hematopoietic stem/progenitor cells (HSPC) holds promise for treating X-linked hyper-IgM Syndrome (HIGM1), but its actual therapeutic potential remains elusive. Here, we developed a one-size-fits-all editing strategy for effective T-cell correction, selection, and depletion and investigated the therapeutic potential of T-cell and HSPC therapies in the HIGM1 mouse model. Edited patients' derived CD4 T cells restored physiologically regulated CD40L expression and contact-dependent B-cell helper function. Adoptive transfer of wild-type T cells into conditioned HIGM1 mice rescued antigen-specific IgG responses and protected mice from a disease-relevant pathogen. We then obtained ~ 25% CD40LG editing in long-term repopulating human HSPC. Transplanting such proportion of wild-type HSPC in HIGM1 mice rescued immune functions similarly to T-cell therapy. Overall, our findings suggest that autologous edited T cells can provide immediate and substantial benefits to HIGM1 patients and position T-cell ahead of HSPC gene therapy because of easier translation, lower safety concerns and potentially comparable clinical benefits.