Hereditary neuromuscular diseases are a group of genetic diseases characterized by an onset of the disease in most cases in childhood, having a steadily progressive course of the pathological process, leading to more rapid disability of patients and having a high mortality rate at the age of 18–20 years.Objective. To study a condition of the intra-organ structure in patients with hereditary muscular atrophy and muscular dystrophy using testing of nerve tissue in the organotypic environment in order to justify the prescription of symptomatic neuroprotective therapy.Materials and methods. Ninety patients with hereditary neuromuscular diseases (spinal muscular atrophy types 1, 2 and 3 [n = 30], Duchenne muscular dystrophy [n = 60]) were examined; the control group consisted of 30 healthy people. In vitro – explants of sensory ganglia of 10–12-day-old chicken embryos. A comprehensive clinical, laboratory and experimental study was conducted. Concentrations of neurotrophic factors (Brain Growth Factor, Nerve Growth Factor, Ciliary Neurotrophic Factor) were determined by the enzyme immunoassay method in blood plasma samples using RayBiotech kits and in accordance with the manufacturer's instructions. The experimental study included 300 explants of sensory ganglia of 10–12-day chicken embryos cultured in Petri dishes on collagen substrates in a CO2 incubator (Sanyo, Japan) for 3 days at 36.5 °C and 5% CO2. In order to clarify the biochemical mechanisms involved in pathological cascades in patients with hereditary neuromuscular diseases, a test system was developed that included a sequential analysis of the patient's blood plasma in an organotypic tissue culture at a dilution of 1: 70, followed by an addition of reagents to the medium: synthetic nerve growth factor (NGF) (100 pg/ml). Explant cultivation was carried out according to the method developed at Institute for Physiology n.a. I.P. Pavlov (Saint Petersburg, Russia). Visualization of the objects was made using Axiostar Plus microscope (Carl Zeiss, Germany). The resulting images were analyzed with the help of ImageJ software. A morphometric method was used to quantify the growth of explants. The area index (AI) was calculated as the ratio of the area of the explant growth zone to the initial area. AI reference value was 100%.Results. Brain Growth Factor concentration was at the highest level in patients with progressive amyotrophy, while in patients with progressive myodystrophy, the blood concentra tion of this factor was at a level comparable to the control data, and in some patients the concentration of Brain Growth Factor was lower than normal. NGF concentration showed the highest values in the group of patients with progressive amyotrophies. Blood plasma of patients with progressive amyotrophy dose dependently inhibits the growth of neurites of the spinal ganglia, and blood plasma of patients with myodystrophy has a neurite-weakening effect on the growth of neurites. Introduction of synthetic NGF (100 pg/ml) to organotypic tissue culture containing blood plasma of patients with myodystrophy increased the area index value of 114.0 [111.0; 116.0]%; in explants containing blood plasma of patients with progressive amyotrophy, increased growth of neurites was not observed AI = 80.0 [74.5; 83.0]%.Conclusion. The data obtained are indicative of features of neurotrophic regulation in patients with hereditary muscular atrophy and muscular dystrophy, which should be taken into account when conducting symptomatic treatment aimed at stimulating reparative processes in the nervous tissue. We recommend patients with the neurite-weakening effect of blood plasma to have neuroprotective drugs therapy, and in case of patients with neuritis-inhibiting effect on neurites in organotypic culture of nervous tissue we recommend choosing a drug in vitro individually using pharmacological analysis.
The relative frequency of common neuromuscular diagnoses in a reference center
