Novel PLGA-based nanoformulation decreases doxorubicin-induced cardiotoxicity.

Nanotechnology has the potential to provide formulations of antitumor agents with increased selectivity towards cancer tissue thereby decreasing systemic toxicity. This in vivo study evaluated the potential of novel nanoformulation based on poly(lactic-co-glycolic acid) (PLGA) to reduce the cardiotoxic potential of doxorubicin (DOX). In vivo toxicity of PLGADOX was compared with clinically approved non-PEGylated, liposomal nanoformulation of DOX (LipoDOX) and conventional DOX form (ConvDOX). The study was performed using Wistar Han rats of both sexes that were treated intravenously for 28 days with 5 doses of tested substances at intervals of 5 days. Histopathological analyses of heart tissues showed the presence of myofiber necrosis, degeneration processes, myocytolysis, and hemorrhage after treatment with ConvDOX, whereas only myofiber degeneration and hemorrhage were present after the treatment with nanoformulations. All DOX formulations caused an increase in the troponin T with the greatest increase caused by convDOX. qPCR analyses revealed an increase in the expression of inflammatory markers IL-6 and IL-8 after ConvDOX and an increase in IL-8 expression after lipoDOX treatments. The mass spectra imaging (MSI) of heart tissue indicates numerous metabolic and lipidomic changes caused by ConvDOX, while less severe cardiac damages were found after treatment with nanoformulations. In the case of LipoDOX, autophagy and apoptosis were still detectable, whereas PLGADOX induced only detectable mitochondrial toxicity. Cardiotoxic effects were frequently sex-related with the greater risk of cardiotoxicity observed mostly in male rats.

Paraoxonase and arylesterase activity of paraoxonase 1 and oxidative stress parameters in cervical intraepithelial neoplasia.

Introduction: Paraoxonase 1 (PON1) is the enzyme that removes carcinogenic radicals from lipids. The aim of the study was to investigate the differences in PON1 activity and oxidation stress parameters between patients with cervical intraepithelial neoplasia (CIN) and healthy controls. Materials and methods: The study included 65 women with CIN and 109 healthy women. Lipid parameters were determined on Cobas Integra 400 plus (Roche, Mannheim, Germany). Tiols and reduced glutathione (GSH) were determined spectrophotometric using Eliman reagent. Activity of PON1 was assessed with two substrates, paraoxon and phenylacetate by spectrophotometric method. Malondialdehyde (MDA) was determined by high performance liquid chromatography (Shimadzu Corporation, Kyoto, Japan). Mann-Whitney-test, t-test, χ2-test, correlation and logistic regression was used in statistical analysis. P < 0.05 was considered statistically significant. Results: The basal (P = 0.929) and NaCl-stimulated (P = 0.985) PON1 activity and activities standardised on the concentration of high-density lipoprotein (HDL; P = 0.076; P = 0.065, respectively) and apolipoprotein AI (apo AI; P = 0.444; P = 0.499, respectively) as well as PON1 phenotypes (P = 0.842) did not differ significantly between the groups. The PON1 arylesterase activity (53±19 kU/L vs. 77±17 kU/L; P < 0.001) and HDL-standardized activity (37 (28-44) kU/mmol vs. 43 (37-50) kU/mmol; P < 0.001) and apoAI (29±11 kU/g vs. 44±11 kU/g; P < 0.001) was significantly reduced in the CIN group. The concentration of the thiol groups was similar (P = 0.519), of MDA was lower (0.39 (0.27-0.55) µmol/L vs. 0.76 (0.57-1.15) µmol/L; P < 0.001) and of GSH was higher (112.0 (66.0-129.6) µg/mL vs. 53.4 (34.8-134.4) µg/mL; P < 0.001) in the CIN group. Conclusion: Reduced PON1 arylesterase activity, lower MDA and higher GSH concentration were observed in CIN patients.

Effects of epidurally administered dexmedetomidine and dexamethasone on postoperative pain, analgesic requirements, inflammation, and oxidative stress in thoracic surgery.

The aim of this study was to compare the effects of dexmedetomidine and dexamethasone as adjuvants to preoperative epidural administration of local anesthetic (ropivacaine) in thoracic surgery on the postoperative level of pain, use of analgesics, inflammation, and oxidative stress. The study enrolled 42 patients who underwent elective thoracic surgery in a one-year period at the University Hospital Dubrava (Zagreb, Croatia). Based on a computer-generated randomization list the patients were assigned to the dexmedetomidine (n = 18) or dexamethasone (n = 24) group. Postoperatively, patients of dexmedetomidine group reported lower pain (VAS value 1 h post surgery, 3.4 ± 2.7 vs. 5.4 ± 1.8, dexmedetomidine vs. dexamethasone, p < 0.01) and had lower anal-gesic requirements in comparison with dexamethasone group. Thus, dexmedetomidine in comparison with dexamethasone was more efficient in lowering pain and analgesia requirements 24 h after the surgery. On the contrary, dexamethasone had better anti-inflammatory properties (CRP level 24 h post surgery, 131.9 ± 90.7 vs. 26.0 ± 55.2 mg L-1, dexmedetomidine vs. dexamethasone, p < 0.01). Both dexmedetomidine and dexamethasone exhibited antioxidant effects, however, their antioxidant properties should be further explored. The results of this study improve current knowledge of pain control in thoracic surgery.

Effect of Alpha-Lipoic Acid Supplementation on Low-Grade Squamous Intraepithelial Lesions-Double-Blind, Randomized, Placebo-Controlled Trial.

Low-grade squamous intraepithelial lesion (SIL) is a cytologic diagnosis etiologically related to human papilloma virus (HPV) infection that leads to the release of inflammation mediators, the formation of reactive oxygen species (ROS) and decreased levels of antioxidants in tissues, which is why antioxidants might be considered effective against SIL progression. This randomized double-blind placebo-controlled study aimed to investigate the effectiveness of alpha-lipoic acid (ALA) supplementation (600 mg/day) on the regression of low-grade SIL in 100 patients. Low-grade SIL was determined after the cytological screening, colposcopic examination and targeted biopsy and histological confirmation of cytological−colposcopic diagnosis. Inflammation parameters and the presence of HPV were determined by standard laboratory methods. Dietary and lifestyle habits were investigated using a standardized and validated semi-quantitative food questionnaire (FFQ). ALA supplementation significantly reduced the proportion of patients with low-grade cytological abnormalities, in comparison to placebo. Given the obtained level of significance (p < 0.001), the presented results indicate that short-term ALA supplementation shows a clinically significant effect on cervical cytology. Future studies should focus on the use of innovative formulations of ALA that might induce bioavailability and therapeutic efficiency against HPV infection and the investigation of synergistic effects of concurrent dietary/lifestyle modification and ALA supplementation in both low-grade and high-grade SIL.

Toxicity of nanomixtures to human macrophages: Joint action of silver and polystyrene nanoparticles.

Increasing use of nano-enabled products provides many benefits in various industrial processes and medical applications, but it also raises concern about release of nanoparticles (NPs) into the environment and subsequent human exposure. While potential toxicity of individual NPs types has been well described in scientific literature, exposure and health-related effects of nanomixtures has been poorly described. This study aimed to evaluate the combined effect of silver (AgNP) and polystyrene NPs (PSNP) on the human macrophages. AgNP are one of the most commercialized NPs due to efficient antimicrobial activity, while PSNP are ubiquitous in terrestrial and aquatic environments due to plastic pollution and degradation of polystyrene-based products. Differentiated monocytic cell line THP-1 were used as an in vitro model of human macrophages. Multiple aspects of cellular response to AgNP-PSNP nanomixture were analyzed including cell death, induction of apoptosis, oxidative stress response, expression of pro- and anti-inflammatory cytokines, and nanomechanical properties of cells. NPs uptake was visualized by confocal microscopy and quantified using flow cytometry. Results show that nanomixture increased apoptosis and cell death, expression of IL-6, IL-8 and TNFa, oxidative stress and mitochondrial dysfunction in cells compared to AgNP and PSNP applied as single treatments, indicating mixture additive action. Anti-inflammatory cytokines IL1b, IL-4 and IL-10 were not affected by combined exposure compared to single NPs. Visualization of NPs uptake and internalization showed that AgNP and PSNP were localized mostly in cytoplasm, with small fraction of AgNP translocated into cell nuclei, which explain increased number of double-stranded DNA breaks following exposure of cells to AgNPs alone or in the mixture. Study outcomes represent clear warnings on the human co-exposure to AgNP and PSNP that needs to be implemented in risk assessment approaches towards toxic-free environment.

Tamoxifen Ameliorates Cholestatic Liver Fibrosis in Mice: Upregulation of TGFß and IL6 Is a Potential Protective Mechanism.

The available treatments for cholestatic liver fibrosis are limited, and the disease often progresses to liver cirrhosis. Tamoxifen is a selective modulator of estrogen receptors, commonly used in breast cancer therapy. A recent in vitro study showed that tamoxifen deactivates hepatic stellate cells, suggesting its potential as an antifibrotic therapeutic, but its effects in vivo remain poorly investigated. In the present study, we show that tamoxifen protects against the cholestatic fibrosis induced by a diet supplemented with 0.025% 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC). Mice fed with a DDC-supplemented diet for four weeks and treated with tamoxifen developed a significantly milder degree of liver fibrosis than vehicle-treated mice, as evidenced by a lower percentage of Sirius red-stained area (60.4% decrease in stained area in male and 42% decrease in female mice, p < 0.001 and p < 0.01, respectively) and by lower hydroxyproline content. The finding was further confirmed by qPCR analysis, which showed a lower expression of genes for Col1a1, Acta2, Sox9, Pdgf, and Krt19, indicating the inhibitory effect on hepatic stellate cells, collagen production, and biliary duct proliferation. The degree of protection was similar in male and female mice. Tamoxifen per se, injected into standard-diet-fed mice, increased the expression of genes for Il6 (p < 0.01 and p < 0.001 in male and female mice, respectively) and Tgfß (p < 0.01 for both sexes), and had no adverse effects. We showed that tamoxifen sex-independently protects against cholestatic DDC-induced liver fibrosis. The increased expression of Il6 and Tgfß seems to be a plausible protective mechanism that should be the primary focus of further research.

Cytotoxicity of nanomixture: Combined action of silver and plastic nanoparticles on immortalized human lymphocytes.

BACKGROUND: Silver nanoparticles (AgNP) are one of the most commercialized types of nanomaterials, with a wide range of applications owing to their antimicrobial activity. They are particularly important in hospitals and other healthcare settings, where they are used to maintain sterility of surfaces, textiles, catheters, medical implants, and more. However, AgNP can not only harm bacteria, but also damage mammalian cells and tissue. While the potential toxicity of AgNP is an understood risk, there is a lack of data on their toxicity in combination with polymeric materials, especially plastic nanoparticles such as polystyrene nanoparticles (PSNP) that can be released from surfaces of polystyrene devices during their medical use. AIM: This study aimed to investigate combined effect of AgNP and nanoplastics on human immune response. METHODS: Cells were treated with a range of PSNP and AgNP concentrations, either applied alone or in combination. Cytotoxicity, induction of apoptosis, generation of oxidative stress, uptake efficiency, intracellular localization and nanomechanical cell properties were selected as exposure biomarkers. RESULTS: Collected experimental data showed that nanomixture induced oxidative stress, apoptosis and mortality of Jurkat cells stronger than its individual components. Cell treatment with AgNP/PSNP mixture also significantly changed cell mechanical properties, evidenced by reduction of cells' Young Modulus. CONCLUSION: AgNP and PSNP showed additive toxic effects on immortalized human lymphocytes, evidenced by increase in cellular oxidative stress, induction of apoptosis, and reduction of cell stiffness. These results have important implications for using AgNP and PSNP in medical contexts, particularly for long-term medical implants.

Light and heavy ferritin chain expression in the liver and kidneys of Wistar rats: aging, sex differences, and impact of gonadectomy.

Ferritin is the main intracellular storage of iron. Animal studies show that female liver and kidney express more ferritin and accumulate more iron than male. However, no study so far has investigated sex and age differences in light (FtL) and heavy (FtH) ferritin chain expression. To address this, we relied on specific antibodies and immunochemical methods to analyse the expression of both ferritin chains in the liver and kidney of 3-month and 2-year-old male and female Wistar rats. To see how sex hormones may affect expression we also studied adult animals gonadectomised at the age of 10 weeks. FtL and FtH were more expressed in both organs of female rats, while gonadectomy increased the expression in males and decreased it in females, which suggests that it is stimulated by female and inhibited by male steroid hormones. Normal kidney ferritin distribution and change with aging warrant more attention in studies of (patho) physiological and toxicological processes.

Two Needle Passes Achieve Similar Diagnostic Yield Compared to Three Passes Regarding Diagnosis of Solid Pancreatic Lesions in Endoscopic Ultrasound-Guided Fine Needle Aspiration.

Current guidelines advocate 3-4 passes with a fine-needle aspiration (FNA) to achieve high rates of diagnostic samples for malignancy when performing endoscopic ultrasound (EUS)-guided sampling of solid pancreatic lesions, in the absence of on-site cytologic evaluation. The aim of this study is to compare 2 vs. 3 needle passes in EUS-FNA for solid pancreatic lesions in terms of incremental diagnostic yield and to identify factors associated with the procedure's outcome. In this retrospective study, 2 passes of EUS-FNA were found to have similar diagnostic yield compared to 3 passes for the diagnosis of solid pancreatic masses, suggesting that there might be no significant incremental tissue yield when 3 passes are performed.

Food (Matrix) Effects on Bioaccessibility and Intestinal Permeability of Major Olive Antioxidants.

BACKGROUND: olive pomace extract (OPE) is a rich source of health promoting polyphenols (hydroxytyrosol (HTS) and tyrosol (TS)) and can be used as a nutraceutical ingredient of dietary supplements and functional foods. Its adequate bioavailability is a prerequisite for excreting biological activity and can be significantly and specifically affected by different food matrices. METHODS: in order to investigate food effects on polyphenol bioaccessibility, OPE was co-digested with different foods according to internationally harmonized in vitro digestibility method. Impact of particular nutrients on HTS and TS permeability was assessed on Caco-2 cell monolayer. RESULTS: HTS and TS bioaccessibility and transepithelial permeability can be significantly affected by foods (nutrients), especially by casein and certain types of dietary fiber. Those effects are polyphenol-and nutrient-specific and are achieved either through complexation in gastrointestinal lumen and/or through direct effects of nutrients on intestinal monolayer. CONCLUSIONS: obtained results emphasize the significance and complexity of polyphenol interactions within the food matrix and the necessity of individual investigational approaches with respect to particular food/nutrient and interacting phenolic compounds.

Importance of genetic polymorphisms in liver transplantation outcomes.

Although, liver transplantation serves as the only curative treatment for patients with end-stage liver diseases, it is burdened with complications, which affect survival rates. In addition to clinical risk factors, contribution of recipient and donor genetic prognostic markers has been extensively studied in order to reduce the burden and improve the outcomes. Determination of single nucleotide polymorphisms (SNPs) is one of the most important tools in development of personalized transplant approach. To provide a better insight in recent developments, we review the studies published in the last three years that investigated an association of recipient or donor SNPs with most common issues in liver transplantation: Acute cellular rejection, development of new-onset diabetes mellitus and non-alcoholic fatty liver disease, hepatocellular carcinoma recurrence, and tacrolimus concentration variability. Reviewed studies confirmed previously established SNP prognostic factors, such as PNPLA3 rs738409 for non-alcoholic fatty liver disease development, or the role of CYP3A5 rs776746 in tacrolimus concentration variability. They also identified several novel SNPs, with a reasonably strong association, which have the potential to become useful predictors of post-transplant complications. However, as the studies were typically conducted in one center on relatively low-to-moderate number of patients, verification of the results in other centers is warranted to resolve these limitations. Furthermore, of 29 reviewed studies, 28 used gene candidate approach and only one implemented a genome wide association approach. Genome wide association multicentric studies are needed to facilitate the development of personalized transplant medicine.

Insulin-like growth factor 2 binding protein 3 expression on endoscopic ultrasound guided fine needle aspiration specimens in pancreatic ductal adenocarcinoma.

BACKGROUND: Despite numerous investigations, we still do not have a specific marker for pancreatic ductal adenocarcinoma. Only guideline-recommended biomarker for pancreatic ductal adenocarcinoma is the CA19-9, but it is also present in other gastrointestinal diseases. IMP3 is a new potential biomarker that is over-expressed in some cancers. The aims of our study were (1) to assess IMP3 in benign pancreatic lesions and pancreatic cancer, and (2) to estimate its concentrations in localized and advanced pancreatic cancer. PATIENTS AND METHODS: Seventy-five patients with solid pancreatic lesions who underwent EUS-FNA were included. Patients were divided into three groups: benign lesions, cancer localized only on the pancreas, and patients with advanced pancreatic cancer (locally advanced or with distal metastases). Immunoreactivity of IMP3 was assessed on cytological smears sampled by endoscopic ultrasound. RESULTS: IMP3 was expressed in 89% of the patients with pancreatic cancer and not in benign lesions. Stronger expression of IMP3 protein and stage of the pancreatic cancer was statistically significant. IMP3 was expressed in all localized cancers and in 85% of patients with advanced pancreatic cancer. In the subgroup with locally advanced cancer, IMP3 was expressed in 88%, and in 83% of patients in the subgroup with distal metastasis (P = 0.007). In the present study, sensitivity was 89%, specificity 100%, with positive predictive value of 100% and negative predictive value of 63%. CONCLUSION: There is a positive correlation between IMP3 expression and TNM stages of the pancreatic cancer. Higher expression of IMP3 on EUS-FNA specimens can suggest poorer prognosis.

LPS-induced inflammation desensitizes hepatocytes to Fas-induced apoptosis through Stat3 activation-The effect can be reversed by ruxolitinib.

Recent studies have established a concept of tumour necrosis factor-α (TNF-α)/Fas signalling crosstalk, highlighting TNF-α as a critical cytokine in sensitizing hepatocytes to death induced by Fas activation. However, in the exact inflammatory response, besides TNF-α, many other mediators, that might modulate apoptotic response differentially, are released. To resolve the issue, we studied the effects of lipopolysaccharide (LPS), one of the crucial inductors of inflammation in the liver, on apoptotic outcome. We show that LPS-induced inflammation diminishes the sensitivity of hepatocytes to Fas stimulus in vivo at caspase-8 level. Analysis of molecular mechanisms revealed an increased expression of various pro-inflammatory cytokines in non-parenchymal liver cells and hepatocyte-specific increase in Bcl-xL, associated with signal transducer and activator of transcription 3 (Stat3) phosphorylation. Pre-treatment with ruxolitinib, a selective Janus kinase (JAK) 1/2 inhibitor, prevented the LPS-induced Stat3 phosphorylation and restored the sensitivity of hepatocytes to Fas-mediated apoptosis. Furthermore, ruxolitinib pre-treatment diminished the LPS-induced Bcl-xL up-regulation without an inhibitory effect on LPS-induced expression of pro-inflammatory cytokines. In summary, although the reports are showing that the effects of isolated pro-inflammatory mediators, such as TNF-α or neutrophils, are pro-apoptotic, the overall effect of inflammatory milieu on hepatocytes in vivo is Stat3-dependent desensitization to Fas-mediated apoptosis.

Optimization of a fast screening method for the assessment of low molecular weight thiols in human blood and plasma suitable for biomonitoring studies.

An adequate level of low molecular weight thiols (LMW-SH, especially glutathione (GSH)) protects cellular macromolecules against toxic agents, and is used as a sensitive biomarker of exposure to toxic compounds. During sample collection, storage and preparation, non-enzymatic and enzymatic oxidation of LMW-SH can occur leading to analytical inaccuracy. The aim of this study was to optimize a fast and reliable screening method for the determination of LMW-SH, mainly GSH, in blood and plasma samples as well as to investigate the impact of storage conditions on the LMW-SH stability. Based on our results, the described spectrophotometric method allows fast and reliable determination of LMW-SH in blood and plasma samples. Results on incubation of samples at 37 °C imply that synthesis of LMW-SH (probably GSH) as well as dynamic interexchange among various thiols forms can be induced in blood cells in in vitro conditions. Importantly, the level of LMW-SH in blood and plasma stored at -20 °C was constant, indicating that they can be stored at -20 °C for at least 30 days. Therefore, the method is suitable for assessment of LMW-SH in long-term human biomonitoring as well as environmental field studies, especially those involving a large number of samples such as epidemiological studies.

A modern approach to the treatment of plaque psoriasis.

Psoriasis is a common chronic inflammatory skin disease which affects 0.5-1 % of children and 2-3 % of the adult population. In Croatia, 1.6 % of the population suffer from psoriasis. Distribution of the disease is bimodal, with the first peak at the age of 20-30, and the second at the age of 50-60. The etiopathogenesis of the disease is multifactorial, the key factors being genetic predisposition combined with immunological disorders, environmental factors and skin barrier damage. There are several clinical variants of the disease. The main signalling pathways in psoriasis include TNF-α, IL-23 and IL-17. Topical agents are used for the treatment of the mild form, and the systemic conventional therapy is used for the treatment of moderate to severe forms of the disease. In cases where's no response, or intolerance or contraindications are present, new targeted medications are to be administered. Development in the field of immunogenetics of psoriasis leads to personalized medicine.

Targeting Tumor Markers with Antisense Peptides: An Example of Human Prostate Specific Antigen.

The purpose of this paper was to outline the development of short peptide targeting of the human prostate specific antigen (hPSA), and to evaluate its effectiveness in staining PSA in human prostate cancer tissue. The targeting of the hPSA antigen by means of antisense peptide AVRDKVG was designed according to a three-step method involving: 1. The selection of the molecular target (hPSA epitope), 2. the modeling of an antisense peptide (paratope) based on the epitope sequence, and 3. the spectroscopic evaluation of sense-antisense peptide binding. We then modified standard hPSA immunohistochemical staining practice by using a biotinylated antisense peptide instead of the standard monoclonal antibody and compared the results of both procedures. Immunochemical testing on human tissue showed the applicability of the antisense peptide technology to human molecular targets. This methodology represents a new approach to deriving peptide ligands and potential lead compounds for the development of novel diagnostic substances, biopharmaceuticals and vaccines.

Higher Post-Operative Serum Vitamin D Level is Associated with Better Survival Outcome in Colorectal Cancer Patients.

25-Hydroxyvitamin D (25-OHD) may have a prognostic value in colorectal cancer (CRC) patients. However, as 25-OHD concentration is strongly impacted by surgery, it is uncertain what is the most reliable time-point for 25-OHD assessment, pre- or post-operative. Therefore, we examined 515 CRC patients (AJCC I-III) who underwent surgery. Blood samples were collected either pre-operatively (n = 286; median = 1 day before surgery) or post-operatively (n = 229; median = 8 days). Serum 25-OHD concentration was determined by liquid chromatography-tandem mass spectrometry. Association between 25-OHD and survival was tested in the whole cohort, followed by stratified analyses in pre- and post-operatively sampled. Median 25-OHD in the cohort was 36.7 nmol/L and median follow-up time was 5.9 years. There were no differences between pre- and post-operative cohort in age, sex, 25-OHD, AJCC stage, or localization of tumor. After adjustment, higher 25-OHD (>50 nmol/L) was associated with better overall survival only in post-operative (HR = 0.53; 95% CI: 0.33-0.84; P = 0.006), but not in pre-operative cohort (HR = 1.13; 95% CI: 0.77-1.65; P = 0.53). In conclusion, higher post-operative 25-OHD levels were associated with better survival outcome in CRC patients, while no such association was found for pre-operative levels. Time-point of blood collection should be addressed carefully in future research as it might affect the prognostic value of 25-OHD in CRC.

Synergistic effects of parabens and plastic nanoparticles on proliferation of human breast cancer cells.

Many personal care products on the market contain endocrine disrupting chemicals, including parabens. Parabens are well known chemical additives used as preservatives. They have been found in mammary glands and breast cancer tissues. At the same time, the general public is increasingly exposed to plastic micro- and nanoparticles generated during plastic production and waste disposal. Exposure to chemical cocktails is a realistic scenario of high public health interest, in which many types of compounds such as these two may exhibit synergistic or additive adverse effects. This study evaluated the effects of plastic nanoparticles, parabens, and their mixture on the viability and proliferation of two human breast cancer cell lines: MDA-MB 231, which lacks oestrogen receptors, and MCF-7, which expresses these receptors. Parabens increased proliferation of oestrogen-sensitive breast cancer cells, and this effect became synergistic in the presence of plastic nanoparticles. The mechanism behind synergy may be related to the translocation and adsorption properties of nanoplastics, which served as a Trojan horse to expose cells to parabens more efficiently. These preliminary findings support growing evidence warning about the urgent problem of human exposure to combinations of plastic waste and contingent chemicals.

Forgotten partners and function regulators of inducible metallothioneins.

Metallothioneins are peculiar cysteine rich, heat resistant, small cellular plasma proteins expressed through almost all life forms. The currently established biological functions of metallothioneins are the homeostasis of essential metals and protection against toxic transitional metals (TM) alongside defence from oxidative stress by direct scavenging of reactive oxygen and nitrogen species (ROS and RNS). In mammals, among the four main evolutionary conserved forms, only the ubiquitously expressed metallothionein 1 and 2 (here abbreviated as MT) are inducible by TM, oxidative stress, glucocorticoids and starvation among various other stimuli. However, more than sixty years after being discovered, metallothioneins still bear unresolved issues about their possible physiological function and regulation. The biological function of MTs has still not been associated with the in vitro-demonstrated capacity of MT interaction with cellular molecules glutathione (GSH) or adenosine triphosphate (ATP), or with the possibility of direct iron-MT binding in the reducing intracellular environment of some organelles, e.g. lysosomes. Iron as the most abundant cellular TM is also one of the main physiological sources of ROS. Moreover, iron exhibits strain, sex and age differences that reflected ROS generation and MT induction in (patho)physiology and toxicology studies. A recent study showed that iron sex differences follows expression of both ferritin and MT leading to wide implications from essential TM interconnectivity to aging. This review places emphasis on biochemically proven but physiologically ignored interactions of MT with iron to stimulate advanced research for establishing a wide frame of the biological roles of MTs important for health and longevity.

Impact of pharmacotherapeutic education on medication adherence and adverse outcomes in patients with type 2 diabetes mellitus: a prospective, randomized study.

AIM: To evaluate the impact of pharmacotherapeutic education on 30-day post-discharge medication adherence and adverse outcomes in patients with type 2 diabetes mellitus (T2DM). METHODS: The prospective, randomized, single-center study was conducted at the Medical Department of University Hospital Dubrava, Zagreb, between April and June 2018. One hundred and thirty adult patients with T2DM who were discharged to the community were randomly assigned to either the intervention or the control group. Both groups during the hospital stay received the usual diabetes education. The intervention group received additional individual pre-discharge pharmacotherapeutic education about the discharge prescriptions. Medication adherence and occurrence of adverse outcomes (adverse drug reactions, readmission, emergency department visits, and death) were assessed at the follow-up visit, 30 days after discharge. RESULTS: The number of adherent patients was significantly higher in the intervention group (57/64 [89.9%] vs 41/61 [67.2%]; χ2 test, P=0.003]. There was no significant difference between the groups in the number of patients who experienced adverse outcomes (31/64 [48.4%] vs 36/61 [59.0%]; χ2 test, P=0.236). However, higher frequencies of all adverse outcomes were consistently observed in the control group. CONCLUSION: Pharmacotherapeutic education of patients with T2DM can significantly improve 30-day post-discharge medication adherence, without a significant reduction in adverse clinical outcomes. ClinicalTrial.gov identification number: NCT03438162.