Systemic exosomal siRNA delivery reduced alpha-synuclein aggregates in brains of transgenic mice.

Alpha-synuclein (α-Syn) aggregates are the main component of Lewy bodies, which are the characteristic pathological feature in Parkinson's disease (PD) brain. Evidence that α-Syn aggregation can be propagated between neurones has led to the suggestion that this mechanism is responsible for the stepwise progression of PD pathology. Decreasing α-Syn expression is predicted to attenuate this process and is thus an attractive approach to delay or halt PD progression. We have used α-Syn small interfering RNA (siRNA) to reduce total and aggregated α-Syn levels in mouse brains. To achieve widespread delivery of siRNAs to the brain we have peripherally injected modified exosomes expressing Ravies virus glycoprotein loaded with siRNA. Normal mice were analyzed 3 or 7 days after injection. To evaluate whether this approach can decrease α-Syn aggregates, we repeated the treatment using transgenic mice expressing the human phosphorylation-mimic S129D α-Syn, which exhibits aggregation. In normal mice we detected significantly reduced α-Syn messenger RNA (mRNA) and protein levels throughout the brain 3 and 7 days after treatment with RVG-exosomes loaded with siRNA to α-Syn. In S129D α-Syn transgenic mice we found a decreased α-Syn mRNA and protein levels throughout the brain 7 days after injection. This resulted in significant reductions in intraneuronal protein aggregates, including in dopaminergic neurones of the substantia nigra. This study highlights the therapeutic potential of RVG-exosome delivery of siRNA to delay and reverse brain α-Syn pathological conditions

The Role of Inbreeding in the Extinction of a European Royal Dynasty

The kings of the Spanish Habsburg dynasty (1516–1700) frequently married close relatives in such a way that uncle-niece, first cousins and other consanguineous unions were prevalent in that dynasty. In the historical literature, it has been suggested that inbreeding was a major cause responsible for the extinction of the dynasty when the king Charles II, physically and mentally disabled, died in 1700 and no children were born from his two marriages, but this hypothesis has not been examined from a genetic perspective. In this article, this hypothesis is checked by computing the inbreeding coefficient (F) of the Spanish Habsburg kings from an extended pedigree up to 16 generations in depth and involving more than 3,000 individuals. The inbreeding coefficient of the Spanish Habsburg kings increased strongly along generations from 0.025 for king Philip I, the founder of the dynasty, to 0.254 for Charles II and several members of the dynasty had inbreeding coefficients higher than 0.20. In addition to inbreeding due to unions between close relatives, ancestral inbreeding from multiple remote ancestors makes a substantial contribution to the inbreeding coefficient of most kings. A statistically significant inbreeding depression for survival to 10 years is detected in the progenies of the Spanish Habsburg kings. The results indicate that inbreeding at the level of first cousin (F = 0.0625) exerted an adverse effect on survival of 17.8%±12.3. It is speculated that the simultaneous occurrence in Charles II (F = 0.254) of two different genetic disorders: combined pituitary hormone deficiency and distal renal tubular acidosis, determined by recessive alleles at two unlinked loci, could explain most of the complex clinical profile of this king, including his impotence/infertility which in last instance led to the extinction of the dynasty

Expression of a barley cystatin gene in maize enhances resistance against phytophagous mites by altering their cysteine-proteases

Phytocystatins are inhibitors of cysteine-proteases from plants putatively involved in plant defence based on their capability of inhibit heterologous enzymes. We have previously characterised the whole cystatin gene family members from barley (HvCPI-1 to HvCPI-13). The aim of this study was to assess the effects of barley cystatins on two phytophagous spider mites, Tetranychus urticae and Brevipalpus chilensis. The determination of proteolytic activity profile in both mite species showed the presence of the cysteine-proteases, putative targets of cystatins, among other enzymatic activities. All barley cystatins, except HvCPI-1 and HvCPI-7, inhibited in vitro mite cathepsin L- and/or cathepsin B-like activities, HvCPI-6 being the strongest inhibitor for both mite species. Transgenic maize plants expressing HvCPI-6 protein were generated and the functional integrity of the cystatin transgene was confirmed by in vitro inhibitory effect observed against T. urticae and B. chilensis protein extracts. Feeding experiments impaired on transgenic lines performed with T. urticae impaired mite development and reproductive performance. Besides, a significant reduction of cathepsin L-like and/or cathepsin B-like activities was observed when the spider mite fed on maize plants expressing HvCPI-6 cystatin. These findings reveal the potential of barley cystatins as acaricide proteins to protect plants against two important mite pests

Mortality and respiratory support in X-linked myotubular myopathy: a RECENSUS retrospective analysis

PURPOSE: Individuals with X-linked myotubular myopathy (XLMTM) who survive infancy require extensive supportive care, including ventilator assistance, wheelchairs and feeding tubes. Half die before 18 months of age. We explored respiratory support and associated mortality risk in RECENSUS, particularly among patients ≤5 years old who received respiratory support at birth; this subgroup closely matches patients in the ASPIRO trial of gene therapy for XLMTM. // DESIGN: RECENSUS is an international, retrospective study of patients with XLMTM. Descriptive and time-to-event analyses examined survival on the basis of age, respiratory support, tracheostomy use, predicted mutational effects and life-sustaining care. // RESULTS: Outcomes for 145 patients were evaluated. Among 126 patients with respiratory support at birth, mortality was 47% overall and 59% among those ≤5 years old. Median survival time was shorter for patients ≤5 years old than for those >5 years old (2.2 years (IQR 0.7-5.6) vs 30.2 years (IQR 19.4-30.2)). The most common cause of death was respiratory failure (66.7%). Median survival time was longer for patients with a tracheostomy than for those without (22.8 years (IQR 8.7-30.2) vs 1.8 years (IQR 0.2-not estimable)). The proportion of patients living without a tracheostomy was 50% at age 6 months and 28% at age 2 years. Median survival time was longer with provision of life-sustaining care than without (19.4 years (IQR 3.1-not estimable) vs 0.2 years (IQR 0.1-2.1)). CONCLUSIONS: High mortality, principally due to respiratory failure, among patients with XLMTM ≤5 years old despite respiratory support underscores the need for early diagnosis, informed decision-making and disease-modifying therapies

Annexin A6 Is Critical to Maintain Glucose Homeostasis and Survival During Liver Regeneration in Mice

Background and Aims: Liver regeneration requires the organized and sequential activation of events that lead to restoration of hepatic mass. During this process, other vital liver functions need to be preserved, such as maintenance of blood glucose homeostasis, balancing the degradation of hepatic glycogen stores, and gluconeogenesis (GNG). Under metabolic stress, alanine is the main hepatic gluconeogenic substrate, and its availability is the rate‐limiting step in this pathway. Na+‐coupled neutral amino acid transporters (SNATs) 2 and 4 are believed to facilitate hepatic alanine uptake. In previous studies, we demonstrated that a member of the Ca2+‐dependent phospholipid binding annexins, Annexin A6 (AnxA6), regulates membrane trafficking along endo‐ and exocytic pathways. Yet, although AnxA6 is abundantly expressed in the liver, its function in hepatic physiology remains unknown. In this study, we investigated the potential contribution of AnxA6 in liver regeneration. Approach and Results: Utilizing AnxA6 knockout mice (AnxA6−/−), we challenged liver function after partial hepatectomy (PHx), inducing acute proliferative and metabolic stress. Biochemical and immunofluorescent approaches were used to dissect AnxA6−/− mice liver proliferation and energetic metabolism. Most strikingly, AnxA6−/− mice exhibited low survival after PHx. This was associated with an irreversible and progressive drop of blood glucose levels. Whereas exogenous glucose administration or restoration of hepatic AnxA6 expression rescued AnxA6−/− mice survival after PHx, the sustained hypoglycemia in partially hepatectomized AnxA6−/− mice was the consequence of an impaired alanine‐dependent GNG in AnxA6−/− hepatocytes. Mechanistically, cytoplasmic SNAT4 failed to recycle to the sinusoidal plasma membrane of AnxA6−/− hepatocytes 48 hours after PHx, impairing alanine uptake and, consequently, glucose production. Conclusions: We conclude that the lack of AnxA6 compromises alanine‐dependent GNG and liver regeneration in mice

The influence of statins on the free intracellular calcium concentration in human umbilical vein endothelial cells

BACKGROUND: Statins are cholesterol-lowering drugs that are widely used to reduce the risk of cardiac infarction. Their beneficial clinical effects, however, are not restricted to their influence on cholesterol production. As several studies have shown that they have a potency of relaxing blood vessels. METHODS: We measured the effects of statins on the intracellular free calcium concentration ([Ca(2+)](i)) in human umbilical vein endothelial cells (HUVEC) after acute application and 24-h-preincubation of statins. RESULTS: Incubation of the cells for 24 h with cerivastatin or fluvastatin significantly increased the resting [Ca(2+)](i). For cerivastatin this effect manifested at a concentration of 1 μM. Increase of resting [Ca(2+)](i )in the presence of cerivastatin also occurred when the nitric oxide synthase was inhibited. Transient Ca(2+ )release induced by histamine was not affected. CONCLUSIONS: The increase of resting [Ca(2+)](i )after incubation with cerivastatin or fluvastatin may provide an explanation for the direct effects of statins on the endothelial-dependent vasodilatation and restoration of endothelial activity in vivo

Integrative analysis reveals CD38 as a therapeutic target for plasma cell-rich pre-disease and established rheumatoid arthritis and systemic lupus erythematosus

Figure S2. Daratumumab has no impact on T cells and monocytes ex vivo. (A) Total number of CD3+ T cells in each daratumumab concentration at 72 h post-treatment. (B) Quantification of CD38 MFI on CD3+ T cells at 72 h post-culture with isotype control or daratumumab at indicated concentrations. (C) Total number of CD14+ monocytes in each daratumumab concentration at 72 h post-treatment. (D) Quantification of CD38 MFI on CD14+ monocytes at 72 h post-culture with isotype control or daratumumab at indicated concentrations. Data shown represent four patients with SLE, six with RA and six healthy control donors. (PNG 2127 kb

Mitochondrial impairment increases FL-PINK1 levels by calcium-dependent gene expression.

Mutations of the PTEN-induced kinase 1 (PINK1) gene are a cause of autosomal recessive Parkinson's disease (PD). This gene encodes a mitochondrial serine/threonine kinase, which is partly localized to mitochondria, and has been shown to play a role in protecting neuronal cells from oxidative stress and cell death, perhaps related to its role in mitochondrial dynamics and mitophagy. In this study, we report that increased mitochondrial PINK1 levels observed in human neuroblastoma SH-SY5Y cells after carbonyl cyanide m-chlorophelyhydrazone (CCCP) treatment were due to de novo protein synthesis, and not just increased stabilization of full length PINK1 (FL-PINK1). PINK1 mRNA levels were significantly increased by 4-fold after 24h. FL-PINK1 protein levels at this time point were significantly higher than vehicle-treated, or cells treated with CCCP for 3h, despite mitochondrial content being decreased by 29%. We have also shown that CCCP dissipated the mitochondrial membrane potential (Δψm) and induced entry of extracellular calcium through L/N-type calcium channels. The calcium chelating agent BAPTA-AM impaired the CCCP-induced PINK1 mRNA and protein expression. Furthermore, CCCP treatment activated the transcription factor c-Fos in a calcium-dependent manner. These data indicate that PINK1 expression is significantly increased upon CCCP-induced mitophagy in a calcium-dependent manner. This increase in expression continues after peak Parkin mitochondrial translocation, suggesting a role for PINK1 in mitophagy that is downstream of ubiquitination of mitochondrial substrates. This sensitivity to intracellular calcium levels supports the hypothesis that PINK1 may also play a role in cellular calcium homeostasis and neuroprotection

Acute partial Budd-Chiari syndrome and portal vein thrombosis in cytomegalovirus primary infection: a case report

BACKGROUND: Splanchnic vein thrombosis may complicate inherited thrombotic disorders. Acute cytomegalovirus infection is a rare cause of acquired venous thrombosis in the portal or mesenteric territory, but has never been described extending into a main hepatic vein. CASE PRESENTATION: A 36-year-old immunocompetent woman presented with acute primary cytomegalovirus infection in association with extensive thrombosis in the portal and splenic vein. In addition, a fresh thrombus was evident in the right hepatic vein. A thorough evaluation for a hypercoagulable state was negative. The clinical course, biological evolution, radiological and histological findings were consistent with cytomegalovirus hepatitis complicated by a partial acute Budd-Chiari syndrome and portal thrombosis. Therapeutic anticoagulation was associated with a slow clinical improvement and partial vascular recanalization. CONCLUSION: We described in details a new association between cytomegalovirus infection and acute venous thrombosis both in the portal vein and in the right hepatic vein, realizing a partial Budd-Chiari syndrome. One should be aware that this rare thrombotic event may be complicated by partial venous outflow block