ANALYSIS OF ULTRASOUND FIELD PARAMETERS DURING SONICATION EXPERIMENTS IN VITRO—INFLUENCE OF LABORATORY GLASS AND PLASTICS

Apart from being a powerful medical imaging technique ultrasound can also be used as a therapeutic modality. In vitro sonication experiments performed on cultured cells are one of primary research methods. However present sonication protocols and methods meet many effects influencing the final ultrasound dose experienced by the sonicated samples. The main aim of this study is to assess the influence of laboratory glass and plastics on ultrasound field parameters during in vitro sonication experiments. We performed measurements of ultrasound field parameters (ultrasound intensity and its local distribution) behind commonly used laboratory glass and plastics placed into the far field region of an ultrasound transducer. We tested the influence of several types of culture dishes, culture plates and sample test tubes. Culture dishes reduced ultrasound intensity by tens of percent but did not affect the shape of ultrasound field. 6-well plate reduced ultrasound intensity only by 5 %. Culture plates with well diameter smaller than the diameter of the main lobe of ultrasound beam focus ultrasound energy. Laboratory glass and plastics with curved surface also focus ultrasound energy. We proved that laboratory glass and plastics considerably affect ultrasound field parameters. Thus sonicated samples are exposed to different ultrasound conditions compared to those reported in some of scientific articles. Rest of factors (standing waves formation, streaming, cell mixing, heating and homogeneity of ultrasound field in terms of near and far ultrasound field) affecting the ultrasound field parameters experienced by sonicated samples also need to be studied further

THE EFFECT OF ACETYLSALICYLIC ACID ON ANGIOGENESIS IN VITRO

Angiogenesis, the formation of new blood vessels, is an essential aspect of, among others, embryonic development, wound healing and the female reproductive cycle. It is also necessary for the expansion of tumour masses beyond a minute volume. Acetylsalicylic acid (ASA) is a non-steroidal anti-inflammatory drug with additional antitumour activity. We tested ASA for its ability to inhibit angiogenesis in a simplified angiogenesis model, hASC+HUVEC co cultured in vitro, using immunocytochemical staining with fluorescence-marked antibodies and observation of tubulelike structures and their branching under a fluorescence microscope. We confirmed that ASA is an efficient and useful angiogenesis inhibitor and deserves further attention. We intend using the designed angiogenesis model and the methods described for observing changes in angiogenesis after anti tumour photodynamic therapy (PDT), and also for enhancing PDT efficiency by addition of angiogenesis inhibitors

Vitamin B12 in Leber hereditary optic neuropathy mutation carriers: a prospective cohort study

Background Leber hereditary optic neuropathy (LHON) is the most common mitochondrial disorder, frequently resulting in acute or subacute severe bilateral central vision loss. Vitamin B12 deficiency is also a known cause of optic neuropathy through mitochondrial dysfunction. Here we evaluated the prevalence and clinical significance of vitamin B12 deficiency in a large cohort of LHON patients and asymptomatic mutation carriers from a tertiary referral center. Methods From the Munich LHON prospective cohort study, participants included all LHON patients and asymptomatic LHON mutation carriers, who were recruited between February 2014 and March 2020 and consented to participate. Neurological, general, and ophthalmological examinations were regularly performed, as were laboratory tests. Vitamin B12 deficiency was diagnosed if serum vitamin B12 was below 201 pg/mL, or if 201–339 pg/mL plus low serum holotranscobalamin or elevated serum methylmalonic acid or elevated total plasma homocysteine. Results We analyzed 244 subjects, including 147 symptomatic LHON patients (74% males) and 97 asymptomatic mutation carriers (31% males). Median age at study baseline was 34 years (range 5–82 years). The prevalence of vitamin B12 deficiency was higher for LHON mutation carriers than for the general population in all age categories. This was statistically significant for the LHON mutation carriers under 65 years (21% vs. 5–7%, p = 0.002). While vitamin B12 deficiency prevalence was not statistically different between LHON patients and asymptomatic mutation carriers, its clinical correlates, e.g., macrocytosis and polyneuropathy, were more frequent in the subgroup of LHON patients. Excessive alcohol consumption was a significant predictor of vitamin B12 deficiency (p < 0.05). Conclusions The high prevalence of vitamin B12 deficiency in LHON mutation carriers, both asymptomatic mutation carriers and LHON patients, highlights the need for regular vitamin B12 screening in this population, in order to ensure early treatment, aiming for better outcomes. Our study is not conclusive regarding vitamin B12 deficiency as determinant for disease conversion in LHON, and further research is warranted to disentangle the role of vitamin B12 in the pathophysiology and prognosis of LHON

GENE EXPRESSION PROFILING AFTER ANGIOGENESIS INHIBITOR TREATMENT

The angiogenic process can be summarized as cell activation by a lack of oxygen releases angiogenic molecules that attract inflammatory and endothelial cells and promote their proliferation. Several protein fragments produced by the digestion of the blood-vessel walls intensify the proliferative activity of endothelial cells. Acetyl salicylic acid is often used as an analgesic drug to relieve minor aches and pains, a drug with antitumour activity and an anti-inflammatory medication. In our experiments we propose using the angiogenesis model and photodynamic therapy (PDT) for observing changes in angiogenesis after treatment, and also for increasing the effect of PDT by addition of angiogenesis inhibitors

Peripapillary microcirculation in Leber hereditary optic neuropathy

Purpose In this prospective observational comparative case series, we aimed to study the peripapillary capillary network with spectral‐domain optical coherence tomography angiography (OCT‐A) in Leber hereditary optic neuropathy (LHON). Methods Twelve eyes of six individuals, of these three males (five eyes) after clinical onset of visual impairment were imaged by OCT‐A with scans centred on optic discs. Control group consisted of 6 eyes with no visual impairment. Results The three affected individuals lost vision 6 years (at age 22 years), 2 years and 3 months (at age 26 years) and 1 year and 2 months (at age 30 years) prior to OCT‐A examination. All five affected eyes had alterations in density of the radial peripapillary microvascular network at the level of retinal nerve fibre layer, including an eye of a patient treated with idebenone that underwent almost full recovery (best corrected visual acuity 0.87). Interestingly, the other eye showed normal ocular findings 14 months after onset. Results of OCT‐A examination in this eye were unfortunately inconclusive due to a delineation error. At the level of the ganglion cell layer differences could be also noted, but only in two severely affected individuals. There were no differences between unaffected mutation carriers and control eyes. Conclusion Optical coherence tomography angiography scans confirmed that the peripapillary microvascular network is highly abnormal in eyes manifesting visual impairment due to LHON. These findings support the hypothesis that microangiopathy contributes to the development of vision loss in this mitochondrial disorder

FOTOTOXICKÝ VLIV PORFYRINOVÝCH SENSITIZERŮ A VIDITELNÉHO ZÁŘENÍ NA GRAM-POZITIVNÍ METHICILIN-REZISTENTNÍ KMEN S. AUREUS

The use of antimicrobial photodynamic therapy (aPDT) as a therapeutic modality for the treatment of localized microbial infections represents an developing new field. The emergence of strains resistant to antibiotics has provided the necessary impulse for new drug or technology discoveries to combat these resistant compounds. Although the aPDT is still in infancy, its need is still growing. Like PDT, main components of antimicrobial photodynamic therapy are appropriate light, dye called photosensitizer and created reactive oxygen species. In this article photosensitizers TMPyP and ZnTPPS4 are investigated for antimicrobial photodynamic therapy. We tested these porphyrins on bacterial methicilin – resistant strain MRSA alone and bound in complex created with hp-β-cyclodextrin. The light emitting diodes (414 nm) were used at the doses 0 and 150 J/cm2. Tested concentrations were from 0.78 to 100 μM. This experimental work predicated that TMPyP is very successful compound in aPDT. In contrary to ZnTPPS4 which was efficient for eradication of tested gram-positive bacteria only in higher concentrations

Hyperoside as a UV Photoprotective or Photostimulating Compound—Evaluation of the Effect of UV Radiation with Selected UV-Absorbing Organic Compounds on Skin Cells

Ultraviolet (UV) radiation is a non-ionizing radiation, which has a cytotoxic potential, and it is therefore necessary to protect against it. Human skin is exposed to the longer-wavelength components of UV radiation (UVA and UVB) from the sun. In the present paper, we focused on the study of eight organic UV-absorbing compounds: astragalin, beta-carotene, 2,4-dihydroxybenzophenone, 2-hydroxy-4-methoxybenzophenone, hyperoside, 3-(4-methylbenzylidene)camphor, pachypodol, and trans-urocanic acid, as possible protectives of skin cells against UVA and UVB radiation. Their protective effects on skin cell viability, ROS production, mitochondrial membrane potential, liposomal permeability, and DNA integrity were investigated. Only some of the compounds studied, such as trans-urocanic acid and hyperoside, had a significant effect on the examined hallmarks of UV-induced cell damage. This was also confirmed by an atomic force microscopy study of morphological changes in HaCaT cells or a study conducted on a 3D skin model. In conclusion, hyperoside was found to be a very effective UV-protective compound, especially against UVA radiation. Commonly used sunscreen compounds such as 2,4-dihydroxybenzophenone, 2-hydroxy-4-methoxybenzophenone, and 3-(4-methylbenzylidene)camphor turned out to be only physical UV filters, and pachypodol with a relatively high absorption in the UVA region was shown to be more phototoxic than photoprotective

Study of photodynamic, sonodynamic and antioxidative influence on HeLa cell line

19-28Photodynamic treatment (PDT) in combination with sonodynamic treatment (SDT) can be used as suitable methods to treat malignant and benign diseases or combat resistant bacteria. Both methods affect the production of reactive oxygen species (ROS). On the other hand, antioxidants are useful for cell protection against ROS. This work was aimed to study the effect of PDT and SDT treatments on the HeLa cell line using antioxidant Pronalen Sensitive Skin® as a protection from free radicals in the cells. We evaluated the effect of sensitizer ClAlPcS2 using battery of in vitro methods, including MTT assay, kinetic production of ROS, mitochondrial membrane potential change, type of cell death and microscopic analysis. Ultrasound treatment was observed to increase the production of ROS, only in combination with PDT, particularly at higher concentrations of ClAlPcS2. The added antioxidant acts as protection against free radicals and has potential as a dietary supplement against aging or free radicals. The results of study suggested that ClAlPcS2 could be used as a potential photosensitizer for treatment of a specific type of cancers