Anti-Melanoma Effects of Miconazole: Investigating the Mitochondria Involvement.

Miconazole is an antimycotic drug showing anti-cancer effects in several cancers. However, little is known on its effects in melanoma. A375 and SK-MEL-28 human melanoma cell lines were exposed to miconazole and clotrimazole (up to 100 mM). Proliferation, viability with MTT assay and vascular mimicry were assayed at 24 h treatment. Molecular effects were measured at 6 h, namely, ATP-, ROS-release and mitochondria-related cytofluorescence. A metabolomic profile was also investigated at 6 h treatment. Carnitine was one of the most affected metabolites; therefore, the expression of 29 genes involved in carnitine metabolism was investigated in the public platform GEPIA2 on 461 melanoma patients and 558 controls. After 24 h treatments, miconazole and clotrimazole strongly and significantly inhibited proliferation in the presence of 10% serum on either melanoma cell lines; they also strongly reduced viability and vascular mimicry. After 6 h treatment, ATP reduction and ROS increase were observed, as well as a significant reduction in mitochondria-related fluorescence. Further, in A375, miconazole strongly and significantly altered expression of several metabolites including carnitines, phosphatidyl-cholines, all amino acids and several other small molecules, mostly metabolized in mitochondria. The expression of 12 genes involved in carnitine metabolism was found significantly modified in melanoma patients, 6 showing a significant impact on patients' survival. Finally, miconazole antiproliferation activity on A375 was found completely abrogated in the presence of carnitine, supporting a specific role of carnitine in melanoma protection toward miconazole effect, and was significantly reversed in the presence of caspases inhibitors such as ZVAD-FMK and Ac-DEVD-CHO, and a clear pro-apoptotic effect was observed in miconazole-treated cells, by FACS analysis of Annexin V-FITC stained cells. Miconazole strongly affects proliferation and other biological features in two human melanoma cell lines, as well as mitochondria-related functions such as ATP- and ROS-release, and the expression of several metabolites is largely dependent on mitochondria function. Miconazole, likely acting via carnitine and mitochondria-dependent apoptosis, is therefore suggested as a candidate for further investigations in melanoma treatments.

Implementing Nexobrid for Burn Debridement in a Major Italian Burn Center: A Retrospective Review of Patient Outcomes.

This retrospective study examines the implementation of Nexobrid, an enzymatic debriding agent developed from bromelain, for burn debridement in a major Italian burn center. With previous research showing encouraging results for Nexobrid in terms of reducing the need for surgical intervention and faster eschar removal, the current study aims to add to the growing body of evidence regarding its potential benefits and limitations. The patient database was utilized to identify patients who received Nexobrid treatment between October 2019 and June 2023. A retrospective analysis was conducted to gather demographic information, burn causes, procedural details, and patient outcomes. Of the 30 patients treated with Nexobrid, 10% did not require further surgical intervention, showcasing Nexobrid's potential to improve patient outcomes. However, over 80% of patients still required additional surgical intervention, demonstrating that Nexobrid's effectiveness varies across patients and should be considered a tool rather than a definitive solution in burn wound management. A few patients developed complications, and about 10% of patients succumbed to systemic complications. The study results reveal both the potential benefits and limitations of using Nexobrid in burn debridement. While it successfully eliminated the need for further surgical intervention in a small percentage of patients, the majority still required additional surgical procedures. These findings not only highlight Nexobrid's role as a tool in burn wound management but also point toward the discrepancies with previous studies. The authors suggest future research should include randomized controlled trials, direct comparisons between Nexobrid and traditional debridement methods, and studies incorporating larger and more diverse patient groups.

Nucleic Acid-Sensing Toll-Like Receptors Play a Dominant Role in Innate Immune Recognition of Pneumococci.

Streptococcus pneumoniae (or pneumococcus) is a highly prevalent human pathogen. Toll-like receptors (TLRs) function as immune sensors that can trigger host defenses against this bacterium. Defects in TLR-activated signaling pathways, including deficiency in the adaptor protein myeloid differentiation factor 88 (MyD88), are associated with markedly increased susceptibility to infection. However, the individual MyD88-dependent TLRs predominantly involved in antipneumococcal defenses have not been identified yet. Here we find that triple knockout mice simultaneously lacking TLR7, TLR9, and TLR13, which sense the presence of bacterial DNA (TLR9) and RNA (TLR7 and TLR13) in the phagolysosomes of phagocytic cells, display a phenotype that largely resembles that of MyD88-deficient mice and rapidly succumb to pneumococcal pneumonitis due to defective neutrophil influx into the lung. Accordingly, TLR7/9/13 triple knockout resident alveolar macrophages were largely unable to respond to pneumococci with the production of neutrophil-attracting chemokines and cytokines. Mice with single deficiencies of TLR7, TLR9, or TLR13 showed unaltered ability to control lung infection but were moderately more susceptible to encephalitis, in association with a decreased ability of microglia to mount cytokine responses in vitro Our data point to a dominant, tissue-specific role of nucleic acid-sensing pathways in innate immune recognition of S. pneumoniae and also show that endosomal TLRs are largely capable of compensating for the absence of each other, which seems crucial to prevent pneumococci from escaping immune recognition. These results may be useful to develop novel strategies to treat infections by antibiotic-resistant pneumococci based on stimulation of the innate immune system.IMPORTANCE The pneumococcus is a bacterium that frequently causes infections in the lungs, ears, sinus cavities, and meninges. During these infections, body defenses are triggered by tissue-resident cells that use specialized receptors, such as Toll-like receptors (TLRs), to sense the presence of bacteria. We show here that pneumococci are predominantly detected by TLRs that are located inside intracellular vacuoles, including endosomes, where these receptors can sense the presence of nucleic acids released from ingested bacteria. Mice that simultaneously lacked three of these receptors (specifically, TLR7, TLR9, and TLR13) were extremely susceptible to lung infection and rapidly died after inhalation of pneumococci. Moreover, tissue-resident macrophages from these mice were impaired in their ability to respond to the presence of pneumococci by producing inflammatory mediators capable of recruiting polymorphonuclear leucocytes to infection sites. This information may be useful to develop drugs to treat pneumococcal infections, particularly those caused by antibiotic-resistant strains.

High prevalence of non-HFE gene-associated haemochromatosis in patients from southern Italy.

Hereditary haemochromatosis is an autosomal recessive disorder of iron regulation that results in abnormal intestinal iron absorption with progressive iron overloading of parenchymal cells. Two specific, single point mutations of the HFE gene (C282Y and H63D) have been described in haemochromatosis patients. Epidemiological studies have revealed a strict association between hereditary haemochromatosis and C282Y homozygosis or C282Y/H63D compound heterozygosis, suggesting that these mutations may provide a useful tool for diagnosis. However, recent investigations from southern Europe have reported lower allelic frequencies of the C282Y mutation among haemochromatosis patients, apparently depending on the geographical area of the population analysed. To assess the predictive value of the detection of the C282Y and H63D HFE mutations in our geographical area, we have evaluated their occurrence in 46 haemochromatosis patients from southern Italy. We found that only 19.6% of our patients were homozygous for the C282Y mutation and 21.7% were compound C282Y/H63D heterozygotes. Among the remaining 59%, approximately 40% did not display any of the known HFE mutations. We conclude that, in southern Italy, another genetic determinant/s must be responsible for many haemochromatosis cases and that a genetic screening for the C282Y and H63D HFE mutations is not sufficient for hereditary haemochromatosis diagnosis.

Islet transplantation under the kidney capsule corrects the defects in glycogen metabolism in both liver and muscle of streptozocin-diabetic rats.

Insulin-deficient rats are characterized by multiple defects in the pathway of glycogen synthesis and breakdown in both liver and skeletal muscle. The aim of this study was to clarify whether islet transplantation under the kidney capsule, which is associated with delivery of insulin into the peripheral circulation, is able to normalize glycogen metabolism in liver and muscle of streptozotocin-diabetic rats. Three groups of male Lewis rats were studied under fasting condition: controls, untreated diabetics, and islet transplanted diabetics. Glycogen content, glucose-6-phosphate concentration, and glycogen synthase activity were measured in both liver and skeletal muscle. Untreated diabetic rats were characterized by an increase in glycogen content of 178% and a reduction of glucose-6-phosphate level of 50%. Both glycogen and glucose-6-phosphate contents were restored to normal in transplanted diabetic rats. Active glycogen synthase (0.35 +/- 0.1 nmol/min/mg) and activity ratio (0.22 +/- 0.04) were significantly impaired compared with controls (0.99 +/- 0.2 nmol/min/mg and 0.43 +/- 0.06, respectively) and were normalized by islet transplantation. In the skeletal muscle, glycogen content was similar in the three groups of animals, whereas muscle glucose-6-phosphate level was reduced by 28% and glycogen synthase was in a less active state in the untreated diabetic rats. Both the glucose-6-phosphate concentration and the kinetic profile of glycogen synthase were normalized by islet transplantation. In conclusion, islet transplantation under the kidney capsule corrects the diabetes-induced abnormalities in glycogen and glucose-6-phosphate content and glycogen synthase activity in both liver and skeletal muscle.