Passive transfer of blood sera from ALS patients with identified mutations results in elevated motoneuronal calcium level and loss of motor neurons in the spinal cord of mice

Introduction: Previously, we demonstrated the degeneration of axon terminals in mice after repeated injections of blood sera from amyotrophic lateral sclerosis (ALS) patients with identified mutations. However, whether a similar treatment affects the cell body of motor neurons (MNs) remained unresolved. Methods: Sera from healthy individuals or ALS patients with a mutation in different ALS-related genes were intraperitoneally injected into ten-week-old male Balb/c mice (n = 3/serum) for two days. Afterward, the perikaryal calcium level was measured using electron microscopy. Furthermore, the optical disector method was used to evaluate the number of lumbar MNs. Results: The cytoplasmic calcium level of the lumbar MNs of the ALS-serum-treated mice, compared to untreated and healthy-serum-treated controls, was significantly elevated. While injections of the healthy serum did not reduce the number of MNs compared to the untreated control group, ALS sera induced a remarkable loss of MNs. Discussion: Similarly to the distant motor axon terminals, the injection of blood sera of ALS patients has a rapid degenerative effect on MNs. Analogously, the magnitude of the evoked changes was specific to the type of mutation; furthermore, the degeneration was most pronounced in the group treated with sera from ALS patients with a mutation in the chromosome 9 open reading frame 72 gene

Passive transfer of sera from als patients with identified mutations evokes an increased synaptic vesicle number and elevation of calcium levels in motor axon terminals, similar to sera from sporadic patients

Previously, we demonstrated increased calcium levels and synaptic vesicle densities in the motor axon terminals (MATs) of sporadic amyotrophic lateral sclerosis (ALS) patients. Such alterations could be conferred to mice with an intraperitoneal injection of sera from these patients or with purified immunoglobulin G. Later, we confirmed the presence of similar alterations in the superoxide dismutase 1 G93A transgenic mouse strain model of familial ALS. These consistent observations suggested that calcium plays a central role in the pathomechanism of ALS. This may be further reinforced by completing a similar analytical study of the MATs of ALS patients with identified mutations. However, due to the low yield of muscle biopsy samples containing MATs, and the low incidence of ALS patients with the identified mutations, these examinations are not technically feasible. Alternatively, a passive transfer of sera from ALS patients with known mutations was used, and the MATs of the inoculated mice were tested for alterations in their calcium homeostasis and synaptic activity. Patients with 11 different ALS-related mutations participated in the study. Intraperitoneal injection of sera from these patients on two consecutive days resulted in elevated intracellular calcium levels and increased vesicle densities in the MATs of mice, which is comparable to the effect of the passive transfer from sporadic patients. Our results support the idea that the pathomechanism underlying the identical manifestation of the disease with or without identified mutations is based on a common final pathway, in which increasing calcium levels play a central role

Molecular Characterization of Novel Mycoviruses in Seven Umbelopsis Strains

The presence of viruses is less explored in Mucoromycota as compared to other fungal groups such as Ascomycota and Basidiomycota. Recently, more and more mycoviruses are identified from the early-diverging lineages of fungi. We have determined the genome of 11 novel dsRNA viruses in seven different Umbelopsis strains with next-generation sequencing (NGS). The identified viruses were named Umbelopsis ramanniana virus 5 (UrV5), 6a (UrV6a); 6b (UrV6b); 7 (UrV7); 8a (UrV8a); 8b (UrV8b); Umbelopsis gibberispora virus 1 (UgV1); 2 (UgV2) and Umbelopsis dimorpha virus 1a (UdV1a), 1b (UdV1b) and 2 (UdV2). All the newly identified viruses contain two open reading frames (ORFs), which putatively encode the coat protein (CP) and the RNA-dependent RNA polymerase (RdRp), respectively. Based on the phylogeny inferred from the RdRp sequences, eight viruses (UrV7, UrV8a, UrV8b, UgV1, UgV2, UdV1a, UdV1b and UdV2) belong to the genus Totivirus, while UrV5, UrV6a and UrV6b are placed into a yet unclassified but well-defined Totiviridae-related group. In UrV5, UgV1, UgV2, UrV8b, UdV1a, UdV2 and UdV1b, ORF2 is predicted to be translated as a fusion protein via a rare +1 (or-2) ribosomal frameshift, which is not characteristic to most members of the Totivirus genus. Virus particles 31 to 32 nm in diameter could be detected in the examined fungal strains by transmission electron microscopy. Through the identification and characterization of new viruses of Mucoromycota fungi, we can gain insight into the diversity of mycoviruses, as well as into their phylogeny and genome organization

Vektorkonstrukciók Candida albicans protein foszfatáz Z1 pontmutánsok előállításához

Szakdolgozatom témája a CaPPZ1 gén által kódolt Candida albicans protein foszfatáz Z1 (CaPpz1). Ez az enzim gombaspecifikus foszfatáz, amely szabályozza a sejtfal integritását, a kation homeosztázist, a membránpotenciált, az oxidatív stressz választ és a virulenciát. A CaPPZ1 pontmutációjával olyan integrációs Candida vektorokat hoztam létre, melyek segítségével - a CaPPZ1 deléciós Candida mutáns genomjába integrálva - a CaPpz1 enzim az R262L mutáció következtében inaktív formában, illetve a G2A mutáció eredményeként mirisztoilációs hely nélkül termelődik. A létrejövő mutánsok fenotípusából következtethetünk majd a mutációk fiziológiás szerepére.BSc/BABiológia BScD

Gray matter atrophy is primarily related to demyelination of lesions in multiple sclerosis: a diffusion tensor imaging MRI study

Objective: Cortical pathology, periventricular demyelination and lesion formation in multiple sclerosis (MS) are related (hypothesis 1). Factors in the cerebrospinal fluid close to these compartments could possibly drive the parallel processes. Alternatively, the cortical atrophy could be caused by remote axonal transection (hypothesis 2). Since MRI can differentiate between demyelination and axon loss, we used this imaging modality to investigate the correlation between the pattern of diffusion parameter changes in the periventricular- and deep white matter and the grey matter atrophy. Methods: High-resolution T1-weighted, FLAIR and diffusion MRI images were acquired in 52 RRMS patients and 50 healthy, age-matched controls. We used EDSS to estimate the clinical disability. We used Tract Based Spatial Statistics to compare diffusion parameters (fractional anisotropy, mean, axial and radial diffusivity) between groups. We evaluated global brain, white and grey matter atrophy with SIENAX. Averaged, standard diffusion parameters were calculated in four compartment: periventricular lesioned and normal appearing white matter, non-periventricular lesioned and normal appearing white matter. PLS regression was used to identify which diffusion parameter and in which compartment best predicts the brain atrophy and clinical disability. Results: In our diffusion tensor imaging study compared to controls we found extensive alterations of fractional anisotropy, mean and radial diffusivity and smaller changes of axial diffusivity (maximal p-value>0.0002) in patients that suggested demyelination in the lesioned and in the normal appearing white matter. We found significant reduction in total brain, total white and grey matter (patients: 718.764±14.968cm3, 323.237±7.246cm3, 395.527±8.050cm3, controls: 791.772±22.692cm3 355.350±10.929cm3, 436.422±12.011cm3 (mean±SE)), (p<0.015; p<0.0001; p<0.009; respectively) of patients compared to controls. The PLS analysis revealed a combination of demyelination-like diffusion parameters (higher mean and radial diffusivity in patients) in the lesions and in the non-lesioned periventricular white matter, which best predicted the grey matter atrophy (p<0.001). Similarly, EDSS was best predicted by the radial diffusivity of the lesions and the non- lesioned periventricular white matter, but axial diffusivity of the periventricular lesions also contributed significantly (p<0.0001). Interpretation: Our investigation showed that grey matter atrophy and white matter demyelination are related in MS but white matter axonal loss does not significantly contribute to the grey matter pathology