Regulatory effect of bacterial melanin on the isoforms of new superoxide-producing associates from rat tissues in rotenone-induced Parkinson's disease.

According to recent research, selective neuronal vulnerability in Parkinson's disease (PD) results from several phenotypic traits, including calcium-dependent, feed-forward control of mitochondrial respiration leading to elevated reactive oxygen species and cytosolic calcium concentration, an extensive axonal arbor, and a reactive neurotransmitter. Therefore, antioxidant therapy is a promising direction in the treatment of PD. In vitro studies have indicated the survival-promoting activity of bacterial melanin (BM) on midbrain dopaminergic neuron cultures. It has been established that BM has a number of protective and anti-inflammatory properties, so there is a high probability of a protective effect of BM in the early stages of PD. In this study, PD was induced through the unilateral intracerebral administration of rotenone followed by bacterial melanin. Tissues (brain, lungs, and small intestine) from the observed groups underwent isolation and purification to extract isoforms of new thermostable superoxide (О2-)-producing associates between NADPH-containing lipoprotein (NLP) and NADPH oxidase-Nox (NLP-Nox). The optical absorption spectral characteristics, specific amounts, stationary concentration of the produced О2-, and the content of NADPH in the observed associates were determined. The optical absorption spectra of the NLP-Nox isoforms in the visible and UV regions in the experimental groups did not differ from those of the control group. However, compared with the control group, the specific content of the total fractions of NLP-Nox isoforms associated with PD groups was higher, especially in the small intestine. These findings suggest that the described changes may represent a novel mechanism for rotenone-induced PD. Furthermore, bacterial melanin demonstrated antioxidant properties and regulated membrane formation in the brain, lung, and small intestine. This regulation occurred by inhibiting the release of new membrane-bound formations (NLP-Nox associates) from these membranes while simultaneously regulating the steady-state concentration of the formed О2-.

The comparison of biological effects of bacterial and synthetic melanins in neuroblastoma cells.

Melanins belong to a group of pigments of different structure and origin. They can be produced synthetically or isolated from living organisms. A number of studies have reported testing of various melanins in neurological studies providing different outcomes. Because the structure of melanins can have an effect on obtained results in cell toxicity studies, we present here our original study which aimed to compare the biological effects of bacterial melanin (biotechnologically obtained from B. thuringiensis) with that of synthetic melanin in neuroblastoma cells. Both melanins were structurally characterized in detail. After melanin treatment (0-200 µg/mL), cell viability, glutathione levels, cell morphology and respiration were assessed in SH-SY5Y cells. The structural analysis showed that bacterial melanin is more hydrophilic according to the presence of larger number of -OH moieties. After melanin treatment, we found that synthetic melanin at similar dosage caused always larger cell impairment compared to bacterial melanin. In addition, more severe toxic effect of synthetic melanin was found in mitochondria. In general, we conclude that more hydrophilic, bacterial melanin induced lower toxicity in neuroblastoma cells in comparison to synthetic melanin. Our findings can be useable for neuroscientific studies estimating the potential use for study of neuroprotection, neuromodulation or neurotoxicity.

Detection of Ca(2+)-dependent acid phosphatase activity identifies neuronal integrity in damaged rat central nervous system after application of bacterial melanin.

The study aims to confirm the neuroregenerative effects of bacterial melanin (BM) on central nervous system injury using a special staining method based on the detection of Ca(2+)-dependent acid phosphatase activity. Twenty-four rats were randomly assigned to undergo either unilateral destruction of sensorimotor cortex (group I; n = 12) or unilateral rubrospinal tract transection at the cervical level (C3-4) (group II; n = 12). In each group, six rats were randomly selected after surgery to undergo intramuscular injection of BM solution (BM subgroup) and the remaining six rats were intramuscularly injected with saline (saline subgroup). Neurological testing confirmed that BM accelerated the recovery of motor function in rats from both BM and saline subgroups. Two months after surgery, Ca(2+)-dependent acid phosphatase activity detection in combination with Chilingarian's calcium adenoside triphosphate method revealed that BM stimulated the sprouting of fibers and dilated the capillaries in the brain and spinal cord. These results suggest that BM can promote the recovery of motor function of rats with central nervous system injury; and detection of Ca(2+)-dependent acid phosphatase activity is a fast and easy method used to study the regeneration-promoting effects of BM on the injured central nervous system.

Bacterial melanin promotes recovery after sciatic nerve injury in rats.

Bacterial melanin, obtained from the mutant strain of Bacillus Thuringiensis, has been shown to promote recovery after central nervous system injury. It is hypothesized, in this study, that bacterial melanin can promote structural and functional recovery after peripheral nerve injury. Rats subjected to sciatic nerve transection were intramuscularly administered bacterial melanin. The sciatic nerve transected rats that did not receive intramuscular administration of bacterial melanin served as controls. Behavior tests showed that compared to control rats, the time taken for instrumental conditioned reflex recovery was significantly shorter and the ability to keep the balance on the rotating bar was significantly better in bacterial melanin-treated rats. Histomorphological tests showed that bacterial melanin promoted axon regeneration after sciatic nerve injury. These findings suggest that bacterial melanin exhibits neuroprotective effects on injured sciatic nerve, contributes to limb motor function recovery, and therefore can be used for rehabilitation treatment of peripheral nerve injury.

Bacterial melanin crosses the blood-brain barrier in rat experimental model.

BACKGROUND: Bacterial melanin has been proven to stimulate regeneration after CNS lesions. The purpose of this study was to test, whether bacterial melanin can enter the brain via the blood-brain barrier (BBB). METHODS: Bacterial melanin (BM) was radioactively labeled by the iodobead method and used to test the BBB permeability after systemic injection into rats. The unidirectional influx rate from the blood was calculated by multiple-time regression analysis. A subgroup of the animals was co-injected with non-labeled BM to determine if BM has a saturable transport across the BBB. The levels of radioactivity were determined in the serum and tissues. Arterial blood was sampled to obtain the level of I-BM at different time points after injection. After systemic perfusion with saline, animals were decapitated and brain, spinal cord, liver and kidney samples were obtained and homogenized to test the I-BM level. RESULTS: Study results showed that radioactively-labeled bacterial melanin crossed the BBB, was enzymatically stable in blood and in brain parenchyma. Entry to brain was reduced when non-labeled BM was also present. Circulating melanin entered all regions of the CNS but the uptake was higher in lumbar spinal cord, thalamus, hypothalamus and substantia nigra. Liver and kidneys had high uptake rates of BM. CONCLUSIONS: These results show that bacterial melanin has saturable transport across the BBB and selectively targets some CNS regions. Such transport may contribute to the neuroprotective action of bacterial melanin.

Effects of bacterial melanin on motor recovery and regeneration after unilateral destruction of Substantia Nigra pars compacta in rats.

We examined the potential neuroprotective action of bacterial melanin (BM) in rats after unilateral destruction of Substantia Nigra pars compacta (SNc) dopaminergic neurons. 24 rats were initially trained to an instrumental conditioned reflex (ICR) and then subjected to unilateral electrolytic destruction of SNc. Unilateral deficit in balancing hindlimb movements was observed in all rats after the destruction. On the next day after the destruction part of the animals (n=12) was intramuscularly injected with BM solution at the concentration 6 mg/ml (0.17 g/kg). The other 12 operated rats served as a control group. On the second day after the operation the testing of instrumental conditioned reflex was resumed in both groups. Comparison of recovery periods for the ICR in both groups showed that recovery of the reflex and balancing hindlimb movements in melanin treated rats took place in three postoperative testing days, whereas in control group the recovery was not complete after 23 testing days. Electrophysiological study was conducted in 12 intact rats to show the effects of BM on the activity of SNc neurons. The firing rate of neurons was significantly increased by the BM injection. Morpho-histochemical study of brain sections was conducted after the completion of behavioral experiments. In melanin injected rats the study revealed absence of destruction or electrode trace in Substantia Nigra pars compacta of melanin injected rats. BM stimulates regeneration and microcirculation in SNc. Increased electrical activity of SN neurons and regenerative efforts induced by BM accelerate motor recovery after unilateral SNc destruction.