Anti-inflammatory and antiproliferative evaluation of 4ß-cinnamoyloxy,1ß,3α-dihydroxyeudesm-7,8-ene from Verbesina persicifolia and derivatives.

The anti-inflammatory and antiproliferative activities of 4ß-cinnamoyloxy,1ß,3α-dihydroxyeudesm-7,8-ene (1) and of three derivatives, namely diacetate (2), hydrogenate (3) and diacetate hydrogenate (4) were evaluated. All derivatives exert an anti-inflammatory effect significantly lower than that exerted by 1. Otherwise, both the lead compound and 2-4 showed a comparable antiproliferative activity on human tumor cell lines. The investigation of the mechanism of action accountable for cytotoxicity highlighted the capacity to impair mitochondrial functions through two different pathways, depending on chemical structure. In particular, the lead compound 1 and derivative 3 are able to induce mitochondrial permeability transition, while derivatives 2 and 4 inhibit Complex II in the respiratory chain.

Intravenous magnesium sulfate in pulmonary hypertension of the newborn: a systematic review.

OBJECTIVE: Pulmonary hypertension in the newborn (PPHN) is a significant clinical condition characterized by elevated pulmonary artery pressures, leading to serious health consequences. Magnesium sulfate, known for its vasodilatory properties, has been studied for its potential benefits in managing PPHN. This systematic review evaluates the efficacy and safety of magnesium sulfate in neonates with PPHN. MATERIALS AND METHODS: A systematic literature search was conducted on PubMed and Scopus up to March 10, 2024. Studies were included based on predefined Population, Intervention, Comparison, Outcome, Study (PICOS) criteria focusing on pediatric patients with PPHN treated with magnesium sulfate, compared against placebo or other pharmacological interventions. Outcomes of interest included resolution of PPHN, improved oxygenation, and decreased oxygenation index. RESULTS: From a total of 1,233 articles screened, four studies met the inclusion criteria, including three randomized controlled trials and one multicentric retrospective study. The comparisons included nebulized magnesium sulfate, oral sildenafil, and inhaled nitric oxide. The outcomes varied, with none reported consistently across more than two studies, making a meta-analysis unfeasible. Results indicated a potential benefit of magnesium sulfate in improving pulmonary pressures and oxygenation, but the evidence was insufficient to establish definitive conclusions due to the heterogeneity and a limited number of studies. CONCLUSIONS: The limited data suggest that, while magnesium sulfate may have a role in the management of PPHN, it should not replace established therapies. Further research is needed to better define its efficacy and safety profile.

Hearing Loss and Oxidative Stress: A Comprehensive Review.

Hearing loss is a prevalent condition affecting millions of people worldwide. Hearing loss has been linked to oxidative stress as a major factor in its onset and progression. The goal of this thorough analysis is to investigate the connection between oxidative stress and hearing loss, with an emphasis on the underlying mechanisms and possible treatments. The review addressed the many forms of hearing loss, the role of reactive oxygen species (ROS) in causing damage to the cochlea, and the auditory system's antioxidant defensive mechanisms. The review also goes over the available data that support the use of antioxidants and other methods to lessen hearing loss brought on by oxidative stress. We found that oxidative stress is implicated in multiple types of hearing loss, including age-related, noise-induced, and ototoxic hearing impairment. The cochlea's unique anatomical and physiological characteristics, such as high metabolic activity and limited blood supply, make it particularly susceptible to oxidative damage. Antioxidant therapies have shown promising results in both animal models and clinical studies for preventing and mitigating hearing loss. Emerging therapeutic approaches, including targeted drug delivery systems and gene therapy, offer new possibilities for addressing oxidative stress in the auditory system. The significance of this review lies in its comprehensive analysis of the intricate relationship between oxidative stress and hearing loss. By synthesizing current knowledge and identifying gaps in understanding, this review provides valuable insights for both researchers and clinicians. It highlights the potential of antioxidant-based interventions and emphasizes the need for further research into personalized treatment strategies. Our findings on oxidative stress mechanisms may also affect clinical practice and future research directions. This review serves as a foundation for developing novel therapeutic approaches and may inform evidence-based strategies for the prevention and treatment of hearing loss, ultimately contributing to improved quality of life for millions affected by this condition worldwide.

Synthesis and biological activity of 1,4-dihydrobenzothiopyrano[4,3-c]pyrazole derivatives, novel pro-apoptotic mitochondrial targeted agents.

This study reports the synthesis of a number of 1- and 2-phenyl derivatives of the 1,4-dihydrobenzothiopyrano[4,3-c]pyrazole nucleus, which were obtained by the reaction of the versatile 7-substituted 2,3-dihydro-3-hydroxymethylene-4H-1-benzothiopyran-4-ones with hydrazine and substituted phenylhydrazines. The antiproliferative activity of the synthesized compounds was evaluated by an in vitro assay on human tumor cell lines (HL-60 and HeLa) and showed a significant capacity of the 7-methoxy-substituted benzothiopyrano[4,3-c]pyrazoles 3b-d, carrying the pendant phenyl group in the 1-position, to inhibit cell growth. Investigation of the mechanism of action indicated the induction of the mitochondrial permeability transition (MPT) as the molecular event responsible for the inhibition of cell growth. This phenomenon is related to the ability of the test compounds to cause a rapid Ca2+-dependent and cyclosporin A-sensitive collapse of the transmembrane potential (DeltaPsi) and matrix swelling. All this leads to the release of caspase activators, such as cytochrome c (cyt c) and apoptosis-inducing factor (AIF), which trigger the pro-apoptotic pathway leading to DNA fragmentation.

Computed Tomography-Based Biomarker for Longitudinal Assessment of Disease Burden in Pulmonary Tuberculosis.

PURPOSE: Computed tomography (CT) images enable capturing specific manifestations of tuberculosis (TB) that are undetectable using common diagnostic tests, which suffer from limited specificity. In this study, we aimed to automatically quantify the burden of Mycobacterium tuberculosis (Mtb) using biomarkers extracted from x-ray CT images. PROCEDURES: Nine macaques were aerosol-infected with Mtb and treated with various antibiotic cocktails. Chest CT scans were acquired in all animals at specific times independently of disease progression. First, a fully automatic segmentation of the healthy lungs from the acquired chest CT volumes was performed and air-like structures were extracted. Next, unsegmented pulmonary regions corresponding to damaged parenchymal tissue and TB lesions were included. CT biomarkers were extracted by classification of the probability distribution of the intensity of the segmented images into three tissue types: (1) Healthy tissue, parenchyma free from infection; (2) soft diseased tissue, and (3) hard diseased tissue. The probability distribution of tissue intensities was assumed to follow a Gaussian mixture model. The thresholds identifying each region were automatically computed using an expectation-maximization algorithm. RESULTS: The estimated longitudinal course of TB infection shows that subjects that have followed the same antibiotic treatment present a similar response (relative change in the diseased volume) with respect to baseline. More interestingly, the correlation between the diseased volume (soft tissue + hard tissue), which was manually delineated by an expert, and the automatically extracted volume with the proposed method was very strong (R2 ≈ 0.8). CONCLUSIONS: We present a methodology that is suitable for automatic extraction of a radiological biomarker from CT images for TB disease burden. The method could be used to describe the longitudinal evolution of Mtb infection in a clinical trial devoted to the design of new drugs.

Discovery of a new anilino-3H-pyrrolo[3,2-f]quinoline derivative as potential anti-cancer agent.

The newly synthesized 1-[4-(3H-pyrrolo[3,2-f]quinolin-9-ylamino)-phenyl]-ethanone hydrochloride showed high antiproliferative activity by mixed mechanisms of action. The compound acts by forming an intercalative complex with DNA and inhibiting DNA topoisomerase II (topo II) and by blocking the cell cycle in G(2)/M phase. Probable cell death by apoptosis is also suggested by flow cytometry analysis.

Defective positioning in granulomas but not lung-homing limits CD4 T-cell interactions with Mycobacterium tuberculosis-infected macrophages in rhesus macaques.

Protection against Mycobacterium tuberculosis (Mtb) infection requires CD4 T cells to migrate into the lung and interact with infected macrophages. In mice, less-differentiated CXCR3+ CD4 T cells migrate into the lung and suppress growth of Mtb, whereas CX3CR1+ terminally differentiated Th1 cells accumulate in the blood vasculature and do not control pulmonary infection. Here we examine CD4 T-cell differentiation and lung homing during primary Mtb infection of rhesus macaques. Mtb-specific CD4 T cells simultaneously appeared in the airways and blood ∼21-28 days post exposure, indicating that recently primed effectors are quickly recruited into the lungs after entering circulation. Mtb-specific CD4 T cells in granulomas display a tissue-parenchymal CXCR3+CX3CR1-PD-1hiCTLA-4+ phenotype. However, most granuloma CD4 T cells are found within the outer lymphocyte cuff and few localize to the myeloid cell core containing the bacilli. Using the intravascular stain approach, we find essentially all Mtb-specific CD4 T cells in granulomas have extravasated across the vascular endothelium into the parenchyma. Therefore, it is unlikely to be that lung-homing defects introduced by terminal differentiation limit the migration of CD4 T cells into granulomas following primary Mtb infection of macaques. However, intralesional positioning defects within the granuloma may pose a major barrier to T-cell-mediated immunity during tuberculosis.

The extreme sensitivity of Mycobacterium tuberculosis to the front-line antituberculosis drug isoniazid.

Mycobacterium tuberculosis is a natural mutant in oxyR, a close homolog of the central regulator of peroxide stress response in enteric bacteria. Inactivation of oxyR is specific for M. tuberculosis and other members of the M. tuberculosis complex. This phenomenon appears as a paradox due to the ability of this organism to parasitize host macrophages, in which the ingested organisms are likely to be exposed to reactive oxygen intermediates. However, the surprising finding that M. tuberculosis has multiple deletions, nonsense and frameshift mutations in oxyR may help explain the exceptionally high sensitivity of M. tuberculosis to the potent antituberculosis agent isoniazid. One of the genes affected by oxyR lesions, ahpC (encoding an alkylhydroperoxide reductase) may determine the intrinsic sensitivity of mycobacteria to isoniazid.

Interferon-gamma response to the treatment of active pulmonary and extra-pulmonary tuberculosis.

BACKGROUND: Interferon-gamma (IFN-γ) release assays (IGRAs) are used to diagnose tuberculosis (TB) but not to measure treatment response. OBJECTIVE: To measure IFN-γ response to active anti-tuberculosis treatment. DESIGN: Patients from the Henan Provincial Chest Hospital, Henan, China, with TB symptoms and/or signs were enrolled into this prospective, observational cohort study and followed for 6 months of treatment, with blood and sputum samples collected at 0, 2, 4, 6, 8, 16 and 24 weeks. The QuantiFERON® TB-Gold assay was run on collected blood samples. Participants received a follow-up telephone call at 24 months to determine relapse status. RESULTS: Of the 152 TB patients enrolled, 135 were eligible for this analysis: 118 pulmonary (PTB) and 17 extra-pulmonary TB (EPTB) patients. IFN-γ levels declined significantly over time among all patients (P = 0.002), with this decline driven by PTB patients (P = 0.001), largely during the initial 8 weeks of treatment (P = 0.019). IFN-γ levels did not change among EPTB patients over time or against baseline culture or drug resistance status. CONCLUSION: After 6 months of effective anti-tuberculosis treatment, IFN-γ levels decreased significantly in PTB patients, largely over the initial 8 weeks of treatment. IFN-γ concentrations may offer some value for monitoring anti-tuberculosis treatment response among PTB patients.

Bidirectional transport of spermine in rat liver mitochondria.

Further study of the mitochondrial transport of spermine (Toninello et al. (1988) J. Biol. Chem. 263, 19407) shows that, after loading rat liver mitochondria with [14C]spermine and [32P]phosphate, these components are released together into the surrounding medium by adding mersalyl or N-ethylmaleimide. On later addition of dithioerythritol, both are recaptured, but if acetate or nigericin are added instead, only spermine re-enters and there is continued export of phosphate. This bidirectional transport of spermine in and out mitochondria is driven, respectively, by membrane potential and pH gradient at constant protonmotive force. Results using [14C]spermine or [32P]phosphate, in conjunction with the their unlabelled isomers and with or without carbonyl cyanide/p-trifuloromethoxyphenylhydrazone (FCCP) present suggest that there is a continuous energy-dependent efflux-influx cycling of spermine and phosphate.

Spermine binding to liver mitochondria.

Non-equilibrium binding of spermine to mitochondrial membranes is studied in rat liver mitochondria by applying a new thermodynamic treatment of ligand-receptor interactions (Di Noto, V., Dalla Via, L., Toninello, A. and Vidali, M. (1996) Macromol. Theory Simul. 5, 165-181). The presence on mitochondrial membranes of two spermine binding sites, both with monocoordination, is demonstrated. The calculated binding energy is characteristic for weak interactions. The treatment allows also to evaluate the variations of the molar fraction ratio of spermine bound to sites 1 and 2 as function of total bound spermine. The possible role of the two sites is discussed.

Correlation between cytosolic Ca2+ concentration, protein phosphorylation and platelet secretion.

Addition of the calcium-ionophore ionomycin to acetylsalicylate-treated platelets suspended in a low Ca2+ concentration-containing medium (about 0.1 microM), induced a dose-dependent (range 0.25-3 microM) and transient increase in the cytosolic Ca2+ concentration ([Ca2+]c). Less than 10% of the maximal releasable amount of serotonin was secreted at [Ca2+]c lower than 1 microM, whereas secretion was almost maximal at [Ca2+]c higher than 2 microM. In all cases the secretion stopped after about 1 min even if the [Ca2+]c was kept constant by repeated small additions of CaCl2 (25-40 microM). A rapid phosphorylation of pleckstrin (47 kDa) and myosin light chain (20 kDa) was found in all cases, whereas a weak phosphorylation of a 27 kDa protein occurred at [Ca2+]c lower than 1.5 microM. Addition of 0.2 mM CaCl2 to platelets pretreated for 4 min with 0.5-1 microM ionomycin brought about a serotonin secretion remarkably lower than obtained by the simultaneous addition of CaCl2 and ionophore. Platelets suspended in a low calcium-containing medium and exposed to ionomycin showed a major increase in tyrosine phosphorylation of 60 and 72 kDa proteins and a slight increment in tyrosine phosphorylation of 115 and 130 kDa proteins. Subsequent addition of 0.2 mM CaCl2 induced a widespread phosphotyrosine dephosphorylation, particularly evident in the 60 kDa protein identified as p60c-src kinase. The protein kinase inhibitor genistein caused, together with a marked prevention of the protein tyrosine phosphorylation, a remarkable increase in the ionomycin-elicited secretory activity of platelets All together these results indicate that protein kinase C-dependent pleckstrin phosphorylation is a prerequisite of platelet secretion, but that the latter process is apparently regulated by a network of phosphoproteins, in particular the serine/threonine phosphorylation of 27 and 68 kDa proteins and the tyrosine phosphorylation of the p60c-src were found to be associated with a decrease in the secretory activity.

Spermine binding to liver mitochondria deenergized by ruthenium red plus either FCCP or antimycin A.

Thermodynamic analysis of spermine binding to mitochondria treated with ruthenium red and deenergized with either FCCP or antimycin A confirms the presence of two polyamine binding sites, S1 and S2, both with monocoordination, as previously observed in energized mitochondria [Dalla Via et al., Biochim. Biophys. Acta 1284 (1996) 247-252]. Both sites undergo a marked change in binding capacity and binding affinity upon mitochondrial deenergization. This change is most likely responsible for the incomplete or delayed spermine-mediated inhibition of the permeability transition induced in deenergized mitochondria.

Photochemotherapy in the treatment of cancer.

The development of therapies which are selective for tumor tissues is one of the most important goals in anticancer research. Within this framework photochemotherapy can be considered a very promising approach. Its therapeutic effectiveness depends on two connected factors: drug and light. The drug (photosensitizer) is able to exert an antiproliferative effect only after interaction with suitable light. Both the photosensitizing drug and light alone are ineffective at doses used for these treatments. Nowadays, photochemotherapy is used in the treatment of cutaneous T-cell lymphoma and cavitary tumors. In the first case the photosensitizer is a psoralen derivative (P) and long-wavelength ultraviolet radiation (UVA) is used (PUVA therapy). In the second case, the treatment with porphyrins, porphyrin-based and non porphyrin-based photosensitizers is followed by irradiation with 600-1000 nm light (photodynamic therapy, PDT). This review is concerned with PUVA and PDT treatments of cancer. The molecular mechanisms considered accountable for the photochemotherapeutic effects are discussed, the development of new chemical structures aimed at improving the effectiveness and/or overcoming some undesired side effects will also be reported. Moreover, some clinical applications will be described.

New tetracyclic analogues of photochemotherapeutic drugs 5-MOP and 8-MOP: synthesis, DNA interaction, and antiproliferative activity.

The synthesis of new tetrahydrobenzo- and benzopsoralen derivatives carrying at position 5 or 8 of the furocoumarin moiety a methoxy, hydroxy, or dimethylaminopropoxy side chain is reported. The study of their photoantiproliferative activity and ability to induce erythema on guinea pig skin allows us to state that the derivatives carrying the dimethylaminopropoxy side chain exhibit a very interesting photobiological pattern. Indeed, if compared with the lead compounds 5-MOP and 8-MOP, they exert a higher cytotoxic activity devoid of significant skin phototoxicity. Between them, the more interesting appears to be 16, a nonphototoxic compound whose antiproliferative activity on HeLa cells is 2 orders of magnitude higher than that of the reference drug 8-MOP. Photoreaction experiments have revealed that, like classic furocoumarins, A-T is the preferred nucleic base pair in its photobinding. Moreover, the extent of covalent photoaddition to DNA correlates well with the photobiological activity. For this compound a certain effect was also observed in the dark. Evaluation of the ability to induce DNA cleavage in the presence of human topoisomerase II has suggested that this enzyme is probably the target accountable for this effect.

Isolation of restriction fragments from large plasmids recovered from bacteria with multiple plasmids.

A rapid and simple method for isolation of restriction DNA fragments from large plasmids is described. The loss of large plasmids is avoided by restriction endonuclease cleavage in an agarose gel before DNA precipitation. Plasmids were separated in low-melting-point agarose by electrophoresis, the desired plasmid DNA band was cut from the gel and digested with a restriction endonuclease in the agarose. Restriction fragments in agarose were recovered by a modified phenol-extraction, concentrated with 2-butanol and precipitated with ethanol. The procedure simplifies the task of cloning genes from large plasmids, resulting in high yields of restriction fragments from a desired plasmid in a short time.

The effects of methylglyoxal-bis(guanylhydrazone) on spermine binding and transport in liver mitochondria.

This study evaluated the effect of the anticancer drug methylglyoxal-bis(guanylhydrazone) (MGBG) on the binding of the polyamine spermine to the mitochondrial membrane and its transport into the inner compartment of this organelle. Spermine binding was studied by applying a new thermodynamic treatment of ligand-receptor interactions (Di Noto et al., Macromol Theory Simul 5: 165-181, 1996). Results showed that MGBG inhibited the binding of spermine to the site competent for the first step in polyamine transport; the interaction of spermine with this site, termed S1, also mediates the inhibitory effect of the polyamine on the mitochondrial permeability transition (Dalla Via et al., Biochim Biophys Acta 1284: 247-252, 1996). In the presence of 1 mM MGBG, the binding capacity and affinity of this site were reduced by about 2.6-fold; on the contrary, the binding capacity of the S2 site, which is most likely responsible for the internalization of cytoplasmic proteins (see Dalla Via et al., reference cited above), increased by about 1.3-fold, and its binding affinity remained unaffected. MGBG also inhibited the initial rate of spermine transport in a dose-dependent manner by establishing apparently sigmoidal kinetics. Consequently, the total extent of spermine accumulation inside mitochondria was inhibited. This inhibition in transport seems to reflect a conformational change at the level of the channel protein constituting the polyamine transport system, rather than competitive inhibition at the inner active site of the channel, thereby excluding the possibility that the polyamine and drug use the same transport pathway. Furthermore, it is suggested that, in the presence of MGBG, the S2 site is able to participate in residual spermine transport. MGBG also strongly inhibits deltapH-dependent spermine efflux, resulting in a complete block in the bidirectional flux of the polyamine and its sequestration inside the matrix space. The effects of MGBG on spermine accumulation are consistent with in vivo disruption of the regulator of energy metabolism and replication of the mitochondrial genome.

On the mechanism of the spermine-exerted inhibition on alpha-thrombin-induced platelet activation.

Previous reports have shown that various amines inhibited platelet activation, but no definitive conclusions on their action mechanism were drawn. We have further investigated the action of spermine on platelet responses evoked by alpha-thrombin and other agonists. Spermine inhibited in a concentration-dependent manner (1-10 mM), and more efficiently than spermidine and putrescine, the alpha-thrombin-induced (1.5 nM) platelet activation. Spermine added at a concentration that inhibited completely aggregation only partially affected the thrombin-induced increase in cytosolic Ca(2+) concentration, protein phosphorylation, and ATP secretion. The polyamine had little effect on the morphology of resting platelets, as measured by electron microscopy, thrombin hydrolytic activity, and fibrinogen clotting capacity but decreased the thrombin binding to platelets and isolated glycocalicin. Spermine partially inhibited the aggregation elicited by ADP, vasopressin, platelet-activating factor, thrombin receptor-activating peptide, fluoroaluminate, ionomycin, and dioctanoylglycerol but did not affect the cytosolic Ca(2+) increase induced by these agonists. The polyamine bound to both glycocalicin and platelets, and it inhibited the fibrinogen binding to stimulated platelets. The amount of 14C-spermine bound to resting cells decreased in the presence of the glycoprotein GPIb-antibody LJIB1, whereas the polyamine bound to activated platelets, which was higher than that tied to resting cells, was markedly reduced by LJCP8 or decorsin, a GPIIb/IIIa antibody and antagonist-peptide, respectively. These results indicate that spermine specifically inhibits the thrombin binding to GPIb of resting platelets and the fibrinogen binding to GPIIb/IIIa (integrin alpha(IIb)beta(3)) of activated platelets.

Action of antitumoral platinum complexes on in vitro platelet functions.

This report presents a comparison of the effects of cis- and trans-diamminedichloroplatinum complexes on in vitro platelet functions. Pretreatment of platelets with cis-platinum (cisplatin) induced a slow, dose-dependent (0.1-0.45 mM), increase in the cytosolic Ca2+ concentration, pleckstrin (47 kDa) phosphorylation and serotonin secretion, as well as a slight shape modification with emission of a few pseudopodia. All these effects were remarkably increased in platelets exposed to trans-platinum (transplatin). The rise in cytosolic Ca2+ concentration and serotonin secretion evoked by stimulation of platelets with thrombin were not significantly influenced by cellular exposure to cis-platinum, whereas they were enhanced and inhibited, respectively, by exposure to trans-platinum. Trans-platinum also inhibited thrombin-promoted platelet aggregation to a greater extent than the cis-isomer. While the viscosity of platelet rich-plasma tended to decrease in the presence of cis-platinum, it tended to increase in the presence of trans-platinum. Taken together, these results indicate that the effects on platelet functions of the efficacious antitumor complex cis-platinum is rather different from that of the inactive complex trans-platinum. Therefore, the in vitro tests of platelet functions employed in this study might provide an index of antitumor drug toxicity and serve as a preliminary indicator of therapeutic efficacy.