Quantification of Solid Embryonic Cerebellar Graft Volume in a Degenerative Ataxia Model.

Substitution of lost neurons by neurotransplantation would be a possible management of advanced degenerative cerebellar ataxias in which insufficient cerebellar reserve remains. In this study, we examined the volume and structure of solid embryonic cerebellar grafts in adult Lurcher mice, a model of olivocerebellar degeneration, and their healthy littermates. Grafts taken from enhanced green fluorescent protein (EGFP)-positive embryos were injected into the cerebellum of host mice. Two or six months later, the brains were examined histologically. The grafts were identified according to the EGFP fluorescence in frozen sections and their volumes were estimated using the Cavalieri principle. For gross histological evaluation, graft-containing slices were processed using Nissl and hematoxylin-eosin staining. Adjustment of the volume estimation approach suggested that it is reasonable to use all sections without sampling, but that calculation of values for up to 20% of lost section using linear interpolation does not constitute substantial error. Mean graft volume was smaller in Lurchers than in healthy mice when examined 6 months after the transplantation. We observed almost no signs of graft destruction. In some cases, compact grafts disorganized the structure of the host's cerebellar cortex. In Lurchers, the grafts had a limited contact with the host's cerebellum. Also, graft size was of greater variability in Lurchers than in healthy mice. The results are in compliance with our previous findings that Lurcher phenotype-associated factors have a negative effect on graft development. These factors can hypothetically include cerebellar morphology, local tissue milieu, or systemic factors such as immune system abnormalities.

Lipid Nanoparticles Deliver mRNA to the Brain after an Intracerebral Injection.

Neurological disorders are often debilitating conditions with no cure. The majority of current therapies are palliative rather than disease-modifying; therefore, new strategies for treating neurological disorders are greatly needed. mRNA-based therapeutics have great potential for treating such neurological disorders; however, challenges with delivery have limited their clinical potential. Lipid nanoparticles (LNPs) are a promising delivery vector for the brain, given their safer toxicity profile and higher efficacy. Despite this, very little is known about LNP-mediated delivery of mRNA into the brain. Here, we employ MC3-based LNPs and successfully deliver Cre mRNA and Cas9 mRNA/Ai9 sgRNA to the adult Ai9 mouse brain; greater than half of the entire striatum and hippocampus was found to be penetrated along the rostro-caudal axis by direct intracerebral injections of MC3 LNP mRNAs. MC3 LNP Cre mRNA successfully transfected cells in the striatum (∼52% efficiency) and hippocampus (∼49% efficiency). In addition, we demonstrate that MC3 LNP Cas9 mRNA/Ai9 sgRNA edited cells in the striatum (∼7% efficiency) and hippocampus (∼3% efficiency). Further analysis demonstrates that MC3 LNPs mediate mRNA delivery to multiple cell types including neurons, astrocytes, and microglia in the brain. Overall, LNP-based mRNA delivery is effective in brain tissue and shows great promise for treating complex neurological disorders.

Experimental Treatment with Edaravone in a Mouse Model of Spinocerebellar Ataxia 1.

Edaravone is a mitochondrially targeted drug with a suggested capability to modify the course of diverse neurological diseases. Nevertheless, edaravone has not been tested yet in the context of spinocerebellar ataxia 1 (SCA1), an incurable neurodegenerative disease characterized mainly by cerebellar disorder, with a strong contribution of inflammation and mitochondrial dysfunction. This study aimed to address this gap, exploring the potential of edaravone to slow down SCA1 progression in a mouse knock-in SCA1 model. SCA1154Q/2Q and healthy SCA12Q/2Q mice were administered either edaravone or saline daily for more than 13 weeks. The functional impairments were assessed via a wide spectrum of behavioral assays reflecting motor and cognitive deficits and behavioral abnormalities. Moreover, we used high-resolution respirometry to explore mitochondrial function, and immunohistochemical and biochemical tools to assess the magnitude of neurodegeneration, inflammation, and neuroplasticity. Data were analyzed using (hierarchical) Bayesian regression models, combined with the methods of multivariate statistics. Our analysis pointed out various previously documented neurological and behavioral deficits of SCA1 mice. However, we did not detect any plausible therapeutic effect of edaravone on either behavioral dysfunctions or other disease hallmarks in SCA1 mice. Thus, our results did not provide support for the therapeutic potential of edaravone in SCA1.

Consensus Paper: Strengths and Weaknesses of Animal Models of Spinocerebellar Ataxias and Their Clinical Implications.

Spinocerebellar ataxias (SCAs) represent a large group of hereditary degenerative diseases of the nervous system, in particular the cerebellum, and other systems that manifest with a variety of progressive motor, cognitive, and behavioral deficits with the leading symptom of cerebellar ataxia. SCAs often lead to severe impairments of the patient's functioning, quality of life, and life expectancy. For SCAs, there are no proven effective pharmacotherapies that improve the symptoms or substantially delay disease progress, i.e., disease-modifying therapies. To study SCA pathogenesis and potential therapies, animal models have been widely used and are an essential part of pre-clinical research. They mainly include mice, but also other vertebrates and invertebrates. Each animal model has its strengths and weaknesses arising from model animal species, type of genetic manipulation, and similarity to human diseases. The types of murine and non-murine models of SCAs, their contribution to the investigation of SCA pathogenesis, pathological phenotype, and therapeutic approaches including their advantages and disadvantages are reviewed in this paper. There is a consensus among the panel of experts that (1) animal models represent valuable tools to improve our understanding of SCAs and discover and assess novel therapies for this group of neurological disorders characterized by diverse mechanisms and differential degenerative progressions, (2) thorough phenotypic assessment of individual animal models is required for studies addressing therapeutic approaches, (3) comparative studies are needed to bring pre-clinical research closer to clinical trials, and (4) mouse models complement cellular and invertebrate models which remain limited in terms of clinical translation for complex neurological disorders such as SCAs.

[Detection of kidney stones - Imaging and laboratory chemistry including urine analysis]. / Nachweis von Nierensteinen.

Detection of kidney stones - Imaging and laboratory chemistry including urine analysis Abstract. Nephrolithiasis is often the first manifestation of renal colic. This is usually caused by ureteral stones, but also by blood clots, lost papillary necrosis, tumor-related ureteral stenosis or urogenital TB. Today, kidney stones are primarily detected by ultrasound. Although the CT examination without contrast agent is also highly sensitive, because of the radiation exposure, it is only recommended if the findings are unclear. In addition to a careful anamnesis, supplementary diagnostics with urine tests (hematuria and crystalluria, possibly 24-hour urine collection) and serological determinations (creatinine, sodium, potassium, calcium, uric acid, inorganic phosphorus) are important.

Sonic Hedgehog and Triiodothyronine Pathway Interact in Mouse Embryonic Neural Stem Cells.

Neural stem cells are fundamental to development of the central nervous system (CNS)-as well as its plasticity and regeneration-and represent a potential tool for neuro transplantation therapy and research. This study is focused on examination of the proliferation dynamic and fate of embryonic neural stem cells (eNSCs) under differentiating conditions. In this work, we analyzed eNSCs differentiating alone and in the presence of sonic hedgehog (SHH) or triiodothyronine (T3) which play an important role in the development of the CNS. We found that inhibition of the SHH pathway and activation of the T3 pathway increased cellular health and survival of differentiating eNSCs. In addition, T3 was able to increase the expression of the gene for the receptor smoothened (Smo), which is part of the SHH signaling cascade, while SHH increased the expression of the T3 receptor beta gene (Thrb). This might be the reason why the combination of SHH and T3 increased the expression of the thyroxine 5-deiodinase type III gene (Dio3), which inhibits T3 activity, which in turn affects cellular health and proliferation activity of eNSCs.

Two in One: Epithelioid angiomyolipoma within a classic kidney angiomyolipoma - a case report.

BACKGROUND: Epithelioid angiomyolipoma is defined as potentially malignant mesenchymal neoplasm, characterized by proliferating epithelioid cells, whereas classic angiomyolipoma, composed of fat, smooth muscle cells and dysmorphic vessels, is defined as a potentially benign. The usual or classic angiomyolipoma is often found incidentally on imaging studies, relatively easily identified due to the presence of fat, in contrast to the epithelioid angiomyolipoma that can pose diagnostic challenges. CASE PRESENTATION: We report a 51-year-old female patient in which an ultrasonography examination showed a solid mass close to the right renal pelvis with hypoechoic and hyperechoic areas. A differential diagnosis of atypical sinus lipomatosis, lipoma and a transitional cell carcinoma was postulated whereas in a subsequent computed tomography a classic angiomyolipoma was postulated. A re-examination by contrast enhanced ultrasound revealed a striking perfusion difference of the hypoechoic and hyperechoic areas. The hypoechoic area showed homogenous and prolonged enhancement whereas the hypoechoic area displayed a marked slower contrast material flooding and a relatively rapid wash out. The histological analysis from the biopsy of the hyperechoic area showed a classic angiomyolipoma, whereas the sample of the hypoechoic central portion revealed an epithelioid angiomyolipoma. A nephrectomy was performed because of the malignant potential of the epithelioid variant of the angiomyolipoma. CONCLUSIONS: A solid kidney mass with two sharply defined parts, one-part compatible with a classical angiomyolipoma and the other being suspected of carcinoma, is rare, but also illustrative and instructive. The combination of different imaging modalities in the work up of a solid renal mass facilitated to discriminate benign from malignant areas.

Impaired spatial performance in cerebellar-deficient Lurcher mice is not associated with their abnormal stress response.

Both humans and laboratory animals suffering from cerebellar lesions exhibit cognitive as well as many emotional and behavioral abnormalities. These latter have been already observed in the cerebellar mutant mice currently used to highlight some aspect of autism spectrum disorders. The aim of this study was to investigate the influence of cerebellar-related stress response abnormalities on spatial learning and memory. Cerebellar-deficient Lurcher mutant mice were exposed to water environment without active escape possibility and then tested for spatial learning in the Morris water maze. As a marker of stress intensity we measured corticosterone in urine. Finally, the volumes of individual components of the adrenal gland were estimated. Though having spatial navigation deficit in the water maze, Lurcher mice preserved a substantial residuum of learning capacity. Lurcher mutants had a higher increase of corticosterone level after exposure to the water environment than wild type mice. We did not observe any decrease of this physiological stress marker between the start and the end of the spatial navigation task, despite significant improvement of behavioral performances. Furthermore, zona fasciculata and zona reticularis of the adrenal cortex as well as the adrenal medulla were larger in Lurcher mice, reflecting high stress reactivity. We conclude that for both genotypes water exposure was a strong stressor and that there was no habituation to the experiment independently to the increasing controllability of the stressor (e.g. ability to find the escape platform). Based on these findings, we suggest that the enhanced stress response to water exposure is not the main factor explaining the spatial deficits in these cerebellar mutant mice.

Transplantation of Embryonic Cerebellar Grafts Improves Gait Parameters in Ataxic Lurcher Mice.

Hereditary cerebellar ataxias are severe diseases for which therapy is currently not sufficiently effective. One of the possible therapeutic approaches could be neurotransplantation. Lurcher mutant mice are a natural model of olivocerebellar degeneration representing a tool to investigate its pathogenesis as well as experimental therapies for hereditary cerebellar ataxias. The effect of intracerebellar transplantation of embryonic cerebellar solid tissue or cell suspension on motor performance in adult Lurcher mutant and healthy wild-type mice was studied. Brain-derived neurotrophic factor level was measured in the graft and adult cerebellar tissue. Gait analysis and rotarod, horizontal wire, and wooden beam tests were carried out 2 or 6 months after the transplantation. Higher level of the brain-derived neurotrophic factor was found in the Lurcher cerebellum than in the embryonic and adult wild-type tissue. A mild improvement of gait parameters was found in graft-treated Lurcher mice. The effect was more marked in cell suspension grafts than in solid transplants and after the longer period than after the short one. Lurcher mice treated with cell suspension and examined 6 months later had a longer hind paw stride (4.11 vs. 3.73 mm, P < 0.05) and higher swing speed for both forepaws (52.46 vs. 32.79 cm/s, P < 0.01) and hind paws (63.46 vs. 43.67 cm/s, P < 0.001) than controls. On the other hand, classical motor tests were not capable of detecting clearly the change in the motor performance. No strong long-lasting negative effect of the transplantation was seen in wild-type mice, suggesting that the treatment has no harmful impact on the healthy cerebellum.

Malignant hemangiosarcoma in a renal allograft: diagnostic difficulties and clinical course after nephrectomy and immunostimulation.

Hemangiosarcomas are rare tumors of endothelial cell origin. To date, only 20 cases of hemangiosarcoma have been described after renal transplantation, occurring mostly in the skin or in a dialysis fistula. We report a primary metastasizing hemangiosarcoma arising from a renal allograft. The patient was treated with transplant nephrectomy, discontinuation of immunosuppression, and immunostimulation with pegylated interferon-α-2a and has now been in complete remission for 3 years.

Acid-degradable lipid nanoparticles enhance the delivery of mRNA.

Lipid nanoparticle (LNP)-mRNA complexes are transforming medicine. However, the medical applications of LNPs are limited by their low endosomal disruption rates, high toxicity and long tissue persistence times. LNPs that rapidly hydrolyse in endosomes (RD-LNPs) could solve the problems limiting LNP-based therapeutics and dramatically expand their applications but have been challenging to synthesize. Here we present an acid-degradable linker termed 'azido-acetal' that hydrolyses in endosomes within minutes and enables the production of RD-LNPs. Acid-degradable lipids composed of polyethylene glycol lipids, anionic lipids and cationic lipids were synthesized with the azido-acetal linker and used to generate RD-LNPs, which significantly improved the performance of LNP-mRNA complexes in vitro and in vivo. Collectively, RD-LNPs delivered mRNA more efficiently to the liver, lung, spleen and brains of mice and to haematopoietic stem and progenitor cells in vitro than conventional LNPs. These experiments demonstrate that engineering LNP hydrolysis rates in vivo has great potential for expanding the medical applications of LNPs.

Maternal infanticide and low maternal ability in cerebellar mutants Lurcher.

OBJECTIVE: One of the common, but less studied deficiencies in mouse models of cerebellar disorders is impaired breeding capacity. Nevertheless, there is no extensive study in Lurcher (Grid2Lc) mice, a model of olivocerebellar degeneration. The aim of this work was to analyze a breeding capacity of these mutants. METHODS: Lurcher females mated with healthy wild type males were compared with two control groups: wild type females mated with wild type males and wild type females mated with Lurcher males. The breeding capacity of Lurcher mice was analyzed using a fertility rate, mating capability and pups survival rate through three consecutive litters. RESULTS: Lurcher dams did not show significantly reduced fertility and mating capability. Nevertheless, their breeding capacity was affected by reduced litter size, maternal infanticide and higher pup mortality during the maternal care period. CONCLUSION: Lurcher mice are fertile and mating capable cerebellar mutants, but their breeding capacity is reduced due to the postpartum behavioral abnormalities. With regard to hyper-reactivity of the hypothalamo-pituitary-adrenal axis followed by behavioral disinhibition during stressful events in Lurcher mutants, we hypothesize that the lower breeding capacity is associated with these endocrine and behavioral abnormalities.

Near-complete adaptation of the PRiMA knockout to the lack of central acetylcholinesterase.

Acetylcholinesterase (AChE) rapidly hydrolyzes acetylcholine. At the neuromuscular junction, AChE is mainly anchored in the extracellular matrix by the collagen Q, whereas in the brain, AChE is tethered by the proline-rich membrane anchor (PRiMA). The AChE-deficient mice, in which AChE has been deleted from all tissues, have severe handicaps. Surprisingly, PRiMA KO mice in which AChE is mostly eliminated from the brain show very few deficits. We now report that most of the changes observed in the brain of AChE-deficient mice, and in particular the high levels of ambient extracellular acetylcholine and the massive decrease of muscarinic receptors, are also observed in the brain of PRiMA KO. However, the two groups of mutants differ in their responses to AChE inhibitors. Since PRiMA-KO mice and AChE-deficient mice have similar low AChE concentrations in the brain but differ in the AChE content of the peripheral nervous system, these results suggest that peripheral nervous system AChE is a major target of AChE inhibitors, and that its absence in AChE- deficient mice is the main cause of the slow development and vulnerability of these mice. At the level of the brain, the adaptation to the absence of AChE is nearly complete.

[CME-Sonography 104: Angiomyolipomas]. / CME-Sonografie 104: Angiomyolipome.

CME-Sonography 104: Angiomyolipomas Abstract. Angiomyolipomas are the most common benign kidney tumors. Approximately 80 % are spontaneously occurring tumors, the majority <1.0 cm (approximately 54 %). These tumors do not grow and are harmless. Tumors between 1.0 and 2.5 cm (approx. 44 %) also very rarely grow and remain harmless. Larger tumors are rarely found (about 2 %). These must be carefully monitored, as tumors >4.0 cm can spontaneously manifest dangerous bleeding. Apart from spontaneous occurrence, there are also angiomyolipomas in the context of the tuberous sclerosis complex (TCS). Such tumors are often larger and combined with renal cysts and renal cell carcinomas. Measuring the of echo intensity is important for diagnosis.

The Coiled-Coil Forming Peptide (KVSALKE)5 Is a Cell Penetrating Peptide that Enhances the Intracellular Delivery of Proteins.

Protein-based therapeutics have the potential to treat a variety of diseases, however, safe and effective methods for delivering them into cells need to be developed before their clinical potential can be realized. Peptide fusions have great potential for improving intracellular delivery of proteins. However, very few peptides have been identified that can increase the intracellular delivery of proteins, and new peptides that can enhance intracellular protein delivery are greatly needed. In this report, the authors demonstrate that the coiled-coil forming peptide (KVSALKE)5 (termed K5) can function as a cell penetrating peptide (CPP), and can also complex other proteins that contain its partner peptide E5. It is shown here that GFP and Cas9 fused to the K5 peptide has dramatically enhanced cell uptake in a variety of cell lines, and is able to edit neurons and astrocytes in the striatum and hippocampus of mice after a direct intracranial injection. Collectively, these studies demonstrate that the coiled-coil forming peptide (KVSALKE)5 is a new class of multifunctional CPPs that has great potential for improving the delivery of proteins into cells and in vivo.

[CME-Sonography 102/Answers: Acute Epigastric Pain]. / CME-Sonografie 102/Antworten: Akute Oberbauchschmerzen.

CME-Sonography 102/Answers: Acute Epigastric Pain Abstract. Acute upper abdominal pain is a common issue in emergency medicine. Next to the anamnesis and laboratory findings, sonography plays a leading role in differentiation. In addition to the direct representation of the respective pathologies (gallstones, duct dilatations, tumors), the clinical-sonographic examination, the so-called palpation under ultrasound view, is also important. In this article we aim to show the way of diagnosing acute epigastric pain.

[CME Sonography 99: Kidney Echo Changes]. / CME-Sonografie 99: Veränderungen der Nierenechogenität.

CME Sonography 99: Kidney Echo Changes Abstract. Normal kidneys have a smooth contour, kidney length 9-12 cm, volume 90-180 ml/1.73 m2 body surface, parenchyma width 13-18 mm and an echogenicity of the cortex which is hypoechoic compared to the liver and spleen, as well as medullary pyramids, which are again hypoechoic compared to the cortex. Digital ultrasound images enable the echogenicity to be measured between 0 (black) and 255 (white). A normal quotient between the echogenicity of the liver and the renal cortex is 1.0-1.4. A normal quotient between the spleen and the renal cortex is >1.0. The normal quotient between the renal cortex and the renal medulla is >1.0. In diffuse renal parenchyma diseases, changes in echogenicity are observed. There are kidneys with increased echogenicity of the cortex and a ratio to liver and spleen of <1.0 and kidneys with hyperechoic renal medulla and the ratio between renal cortex and renal medulla <1.0.

[CME-Sonography 102: Acute Epigastric Pain]. / CME-Sonografie 102: Akute Oberbauchschmerzen.

CME-Sonography 102: Acute Epigastric Pain Abstract. Acute upper abdominal pain is a common issue in emergency medicine. Next to the anamnesis and laboratory findings, sonography plays a leading role in differentiation. In addition to the direct representation of the respective pathologies (gallstones, duct dilatations, tumors), the clinical-sonographic examination, the so-called palpation under ultrasound view, is also important. In this article we aim to show the way of diagnosing acute epigastric pain.

[CME Sonography 99/Answers: Kidney Echo Changes]. / CME-Sonografie 99/Antworten: Veränderungen der Nierenechogenität.

CME Sonography 99/Answers: Kidney Echo Changes Abstract. Normal kidneys have a smooth contour, kidney length 9-12 cm, volume 90-180 ml/1.73 m2 body surface, parenchyma width 13-18 mm and an echogenicity of the cortex which is hypoechoic compared to the liver and spleen, as well as medullary pyramids, which are again hypoechoic compared to the cortex. Digital ultrasound images enable the echogenicity to be measured between 0 (black) and 255 (white). A normal quotient between the echogenicity of the liver and the renal cortex is 1.0-1.4. A normal quotient between the spleen and the renal cortex is> 1.0. The normal quotient between the renal cortex and the renal medulla is> 1.0. In diffuse renal parenchyma diseases, changes in echogenicity are observed. There are kidneys with increased echogenicity of the cortex and a ratio to liver and spleen of <1.0 and kidneys with hyperechoic renal medulla and the ratio between renal cortex and renal medulla <1.0.

[CME Sonography 96: Spleen Changes]. / CME-Sonografie 96: Milzveränderungen.

CME Sonography 96: Spleen Changes Abstract. The spleen is a somewhat neglected organ but there are some changes that must not be neglected. Splenomegaly, caused worldwide most commonly by malaria, can occur in other infections, in portal vein thrombosis or in portal hypertension in the context of liver cirrhosis. An accessory spleen is also often found. Sometimes after splenectomy, small remains are found which may later hypertrophy. Where focal changes are concerned, we differentiate between various forms of spleen cysts, lymphomas, metastases, and benign tumors.