Protection of the brain from viral infections involves the type I IFN (IFN-I) system, defects in which render humans susceptible to herpes simplex encephalitis (HSE). However, excessive cerebral IFN-I levels lead to pathologies, suggesting the need for tight regulation of responses. Based on data from mouse models, human HSE cases, and primary cell culture systems, we showed that microglia and other immune cells undergo apoptosis in the HSV-1–infected brain through a mechanism dependent on the cyclic GMP-AMP synthase/stimulator of interferon genes (cGAS/STING) pathway, but independent of IFN-I. HSV-1 infection of microglia induced cGAS-dependent apoptosis at high viral doses, whereas lower viral doses led to IFN-I responses. Importantly, inhibition of caspase activity prevented microglial cell death and augmented IFN-I responses. Accordingly, HSV-1–infected organotypic brain slices or mice treated with a caspase inhibitor exhibited lower viral load and an improved infection outcome. Collectively, we identify an activation-induced apoptosis program in brain immune cells that downmodulates local immune responses.

In Wilson Disease (WD), copper accumulates in the liver and brain causing disease. Bis-choline tetrathiomolybdate (TTM) is a potent copper chelator that may have a lower risk of inducing paradoxical neurological worsening than conventional therapy for neurologic WD. To better understand the mode of action of TTM, we investigated the effects of TTM on copper absorption and biliary excretion. In a double-blind randomized setting, hepatic ⁶⁴Cu activity after orally administered ⁶⁴Cu was examined by PET/CT in 16 healthy volunteers before and after 7 days of TTM treatment (15 mg/d) or placebo. Oral ⁶⁴Cu was administered 1 h after final TTM dose. Changes in hepatic ⁶⁴Cu activity reflected changes in intestinal ⁶⁴Cu uptake. Additionally, in four WD patients, distribution of ⁶⁴Cu in venous blood, liver, gallbladder, kidney, and brain were followed after i.v. ⁶⁴Cu dosing for up to 68 hours before and after seven days of TTM (15 mg/d), using PET/MRI. Increased gallbladder ⁶⁴Cu activity was taken as evidence of increased biliary ⁶⁴Cu excretion. In healthy subjects TTM reduced intestinal ⁶⁴Cu uptake by 82% 15 hours after the oral ⁶⁴Cu dose. In WD patients, gallbladder ⁶⁴Cu activity was negligible before and after TTM, while TTM effectively retained ⁶⁴Cu in the blood, significantly reduced hepatic ⁶⁴Cu activity at all time points and significantly reduced cerebral ⁶⁴Cu activity 2 h after the intravenous ⁶⁴Cu dose. While we did not show an increase in biliary excretion of ⁶⁴Cu following TTM administration, we demonstrated that TTM effectively inhibited most intestinal ⁶⁴Cu uptake and retained ⁶⁴Cu in the blood stream, limiting ⁶⁴Cu exposure to organs like the liver and the brain. Bis-choline tetrathiomolybdate (TTM) is an investigational copper chelator for treatment of Wilson Disease. In animal studies of Wilson Disease models, TTM has been shown to facilitate biliary copper excretion. In the present human study, TTM surprisingly did not facilitate biliary copper excretion but instead reduced intestinal copper uptake to a clinically significant degree. Our study improves the understanding of human copper metabolism and the mechanism of action of TTM. Data is presented from two clinical trials:Absorption study: EudraCT 2020-005832-31Excretion study: EudraCT 2021-000102-25.

Excess copper causes hepatocyte death in hereditary Wilson's disease (WD). Current WD treatments by copper-binding chelators may gradually reduce copper overload; they fail, however, to bring hepatic copper close to normal physiological levels. Consequently, lifelong daily dose regimens are required to hinder disease progression. This may result in severe issues due to nonadherence or unwanted adverse drug reactions and also due to drug switching and ultimate treatment failures. This study comparatively tested bacteria-derived copper binding agents-methanobactins (MBs)-for efficient liver copper depletion in WD rats as well as their safety and effect duration. Copper chelators were tested in vitro and in vivo in WD rats. Metabolic cage housing allowed the accurate assessment of animal copper balances and long-term experiments related to the determination of minimal treatment phases. We found that copper-binding ARBM101 (previously known as MB-SB2) depletes WD rat liver copper dose dependently via fecal excretion down to normal physiological levels within 8 days, superseding the need for continuous treatment. Consequently, we developed a new treatment consisting of repetitive cycles, each of ∼1 week of ARBM101 applications, followed by months of in-between treatment pauses to ensure a healthy long-term survival in WD rats. ARBM101 safely and efficiently depletes excess liver copper from WD rats, thus allowing for short treatment periods as well as prolonged in-between rest periods.

Immobilization of the lower limbs promotes a catabolic state that reduces muscle mass, whereas physical training promotes an anabolic state that increases muscle mass. Understanding the molecular mechanisms underlying this is of clinical interest, as loss of muscle mass is a major complication to critical illness in humans. To determine the molecular regulation of protein synthesis and degradation during muscle loss and hypertrophy, we examined skeletal muscle biopsies from healthy human subjects after 2 weeks unilateral immobilization of a lower limb and during 6 weeks of physical rehabilitation. We have previously shown that cross-sectional area of the knee muscle-extensors decreased by ∼10% during immobilization and was completely restored during rehabilitation. Here we provide novel data to suggest that autophagy is an important underlying mechanism involved in regulation of muscle mass. Protein expression of MuRF1 and ATROGIN-1 did not change during the study, indicating that the recruitment of substrates to the proteasomes was unaltered. Phosphorylation of mTORat Ser²⁴⁴⁸ did not change during the study, and neither did phosphorylation of the mTORC1 substrates 4EBP1 Thr^37/46 and p70S6K Thr³⁸⁹, suggesting that this pathway does not suppress protein synthesis during muscle wasting. Protein levels of p62 and ULK1 increased during immobilization and returned to baseline levels during rehabilitation. Same pattern was observed for FOXO3a phosphorylation at Ser^318/321, suggesting transcriptional activation during immobilization and inactivation during rehabilitation. To investigate this further, we analyzed mRNA expression of seven autophagy-related genes controlled by FOXO3a. Five of these (p62, LC3B, BECLIN-1, ATG12, and BNIP3) increased during immobilization and returned to baseline during rehabilitation. In conclusion, immobilization of a lower limb increases autophagy-related gene and protein expression in human skeletal muscle in a pattern that mirrors FOXO3a phosphorylation. These findings could imply that FOXO3a dependent transcriptional regulation of autophagy is involved in the regulation of muscle mass in humans. Clinical Trial Registration: The study was approved by the Ethics Committee of Copenhagen (j.no. H-1-2010-016).

Skeletal muscle satellite cells (SCs) are important for muscle repair and hypertrophy in response mechanical stimuli. Neuron-glial antigen 2-positive (NG2⁺) and alkaline phosphatase-positive (ALP⁺) pericytes may provide an alternative source of myogenic progenitors and/or secrete paracrine factors to induce Pax7⁺ SC proliferation and differentiation. The purpose of this study was to investigate NG2⁺ and ALP⁺ cell quantity, as well as SC content and activation, in human skeletal muscle following prolonged concentric (Conc) or eccentric (Ecc) resistance training. Male subjects engaged in unilateral resistance training utilizing isolated Ecc or Conc contractions. After 12 wk, muscle biopsies were analyzed for NG2⁺ and ALP⁺ pericytes, total Pax7⁺ SCs, activated SCs (Pax7⁺MyoD⁺), and differentiating myogenic cells (Pax7⁻ MyoD⁺). NG2⁺ cells localized to CD31⁺ vessels and the majority coexpressed ALP. NG2⁺ pericyte quantity decreased following both Conc and Ecc training ( P < 0.05). ALP⁺ pericyte quantity declined following Conc ( P < 0.05) but not Ecc training. Conversely, total Pax7⁺ SC content was elevated following Conc only ( P < 0.001), while Pax7⁺MyoD⁺ SC content was increased following Conc and Ecc ( P < 0.001). Follow up analyses demonstrated that CD90⁺ and platelet-derived growth factor receptor-α (PDGFRα)⁺ mononuclear cell proliferation was also increased in response to both Conc and Ecc training ( P < 0.01). In summary, resistance training results in a decline in pericyte quantity and an increase in mesenchymal progenitor cell proliferation, and these events likely influence SC pool expansion and increased activation observed posttraining.

Animal models remain essential to understand the fundamental mechanisms occurring in fetal medicine and obstetric diseases, such as intrauterine growth restriction, preeclampsia and gestational diabetes. These vary regarding the employed method used for induction of the disease, and differ in relation to the animal characteristics (size, number of fetuses, placenta barrier type, etc.). While none of these exactly mirrors the human condition, different pregnant animal models (mice, rats, guinea pigs, chinchillas, rabbits, sheep and pigs) are here described with respect to advantages and limitations. The ability to employ noninvasively diagnostics varies among species, specifically for ultrasound and clinical magnetic resonance imaging procedures. Management of feeding, handling, care and anesthesia are particularly important factors in the pregnant animal.