Various neurodegenerative disorders are associated with human NTE/PNPLA6 dysfunction. Mechanisms of neuropathogenesis in these diseases are far from clearly elucidated. Hereditary spastic paraplegia belongs to a type of neurodegeneration associated with NTE/PNLPLA6 and is implicated in neuron death. In this study, we used Drosophila melanogaster to investigate the consequences of neuronal knockdown of swiss cheese (sws)—the evolutionarily conserved ortholog of human NTE/PNPLA6—in vivo. Adult flies with the knockdown show longevity decline, locomotor and memory deficits, severe neurodegeneration progression in the brain, reactive oxygen species level acceleration, mitochondria abnormalities and lipid droplet accumulation. Our results suggest that SWS/NTE/PNPLA6 dysfunction in neurons induces oxidative stress and lipid metabolism alterations, involving mitochondria dynamics and lipid droplet turnover in neurodegeneration pathogenesis. We propose that there is a complex mechanism in neurological diseases such as hereditary spastic paraplegia, which includes a stress reaction, engaging mitochondria, lipid droplets and endoplasmic reticulum interplay.

Glia are crucial for the normal development and functioning of the nervous system in many animals. Insects are widely used for studies of glia genetics and physiology. Drosophila melanogaster surface glia (perineurial and subperineurial) form a blood–brain barrier in the central nervous system and blood–nerve barrier in the peripheral nervous system. Under the subperineurial glia layer, in the cortical region of the central nervous system, cortex glia encapsulate neuronal cell bodies, whilst in the peripheral nervous system, wrapping glia ensheath axons of peripheral nerves. Here, we show that the expression of the evolutionarily conserved swiss cheese gene is important in several types of glia. swiss cheese knockdown in subperineurial glia leads to morphological abnormalities of these cells. We found that the number of subperineurial glia nuclei is reduced under swiss cheese knockdown, possibly due to apoptosis. In addition, the downregulation of swiss cheese in wrapping glia causes a loss of its integrity. We reveal transcriptome changes under swiss cheese knockdown in subperineurial glia and in cortex + wrapping glia and show that the downregulation of swiss cheese in these types of glia provokes reactive oxygen species acceleration. These results are accompanied by a decline in animal mobility measured by the negative geotaxis performance assay.

Neuropathy target esterase (NTE) is a molecular target for the organophosphorus compound-induced delayed neuropathy (OPIDN) and also one of the genetic factors responsible for the development of the hereditary spastic paraplegia (HSP), characterized by axon degeneration of motoneurons causing progressive lower-limb spastic paralysis. Both HSP and OPIDN are characterized by the distal axonopathy. The molecular mechanisms underlying the axonopathy involved in HSP and OPIDN are poorly understood. In order to have a better understanding of the mechanisms that NTE is involved in, we used one of the homologs, human NTE. Swiss cheese (sws) is a Drosophila melanogaster ortholog of NTE with 39% homology. Mutations in sws as it was shown before lead to age-dependent neurodegeneration, structure alteration of glia cells, and reduced insect life span. To study SWS functions, we used the system of the third-instar larval neuromuscular junctions of D. melanogaster. In this study, we show that mutations in sws (sws1 and sws76−1 ) and SWS knockdown alter neuromuscular junction’s morphology and synaptic microtubules organization.

Aim. To investigate sensitivity of the D. melanogaster neurodegenerative mutants from Swiss cheese group to oxidative stress (OS). Methods. Measuring the life-span, OS resistance, level of lipid peroxidation products, and the number of dophaminergic neurons. Results. We have found decreased life-span, increased sensitivity to OS, increased formation of LPP and degeneration of dophaminergic neurons in brain tissue. Conclusions. Neurodegeneration is associated with the free radical oxidation and is accompanied by accumulation of LPP and increased sensitivity to O

Background. Primary immunodeficiency (PID) compromise the immune system, leaving newborns highly vulnerable to infections. Severe combined immunodeficiency (SCID) is the most severe form, characterized by the absence or dysfunction of T and B cells. Without early treatment, most infants with SCID do not survive their first year. In Ukraine, after a successful pilot project, newborn screening for SCID and other types of PID is now part of Advanced Neonatal Screening, using T-cell receptor excision circles (TREC) and kappa-deleting recombination excision circles (KREC) measurements. Since the pilot project used a different method than the current screening program, the purpose of this study was to compare these two methods, evaluating their benefits and downsides, aiming to optimize screening for early, effective treatment. Materials and Methods. In the pilot project method (Method 1), TREC and KREC quantification was performed using a custom real-time PCR assay with melting curve analysis. Method 1 included standards with known TREC and KREC copy numbers, no-template controls (NTCs), and positive controls to ensure reliable results. The method currently employed in the Advanced Neonatal Screening (Method 2) uses the Biocore® SMA/TKID PLUS Diagnostic Kit, a commercial kit, for TREC, KREC and SMN1 quantification via real-time PCR. Measurements for both methods are reported per one million cells. Results and Discussion. While there are differences in the general parameters of DNA extraction, PCR, and result analysis and interpretation, both Method 1 and Method 2 showed a significant difference in Cq values. Despite these differences, both methods demonstrated the capability of inentifying abnormal TREC/KREC values, enabling the detection of SCID and some PID cases. Conclusion. The pilot project demonstrated the effectiveness of TREC/KREC quantification for SCID screening and led to its implementation in Advanced Neonatal Screening in Ukraine. Over 121,000 newborns were tested, confirming six positive cases. Method 1 provides higher precision and versatility, while Method 2 is faster, simpler, and capable of automation but lacks precise quantification. Adding standards to Method 2 could enhance its utility for widespread SCID screening.