In vitro reconstitution reveals a key role of human mitochondrial EXOG in RNA primer processing.

The removal of RNA primers is essential for mitochondrial DNA (mtDNA) replication. Several nucleases have been implicated in RNA primer removal in human mitochondria, however, no conclusive mechanism has been elucidated. Here, we reconstituted minimal in vitro system capable of processing RNA primers into ligatable DNA ends. We show that human 5'-3' exonuclease, EXOG, plays a fundamental role in removal of the RNA primer. EXOG cleaves short and long RNA-containing flaps but also in cooperation with RNase H1, processes non-flap RNA-containing intermediates. Our data indicate that the enzymatic activity of both enzymes is necessary to process non-flap RNA-containing intermediates and that regardless of the pathway, EXOG-mediated RNA cleavage is necessary prior to ligation by DNA Ligase III. We also show that upregulation of EXOG levels in mitochondria increases ligation efficiency of RNA-containing substrates and discover physical interactions, both in vitro and in cellulo, between RNase H1 and EXOG, Pol γA, Pol γB and Lig III but not FEN1, which we demonstrate to be absent from mitochondria of human lung epithelial cells. Together, using human mtDNA replication enzymes, we reconstitute for the first time RNA primer removal reaction and propose a novel model for RNA primer processing in human mitochondria.

The Role of IFITM Proteins in Tick-Borne Encephalitis Virus Infection.

Tick-borne encephalitis virus (TBEV), of the genus Flavivirus, is a causative agent of severe encephalitis in regions of endemicity of northern Asia and central and northern Europe. Interferon-induced transmembrane proteins (IFITMs) are restriction factors that inhibit the replication cycles of numerous viruses, including flaviviruses such as West Nile virus, dengue virus, and Zika virus. Here, we demonstrate the role of IFITM1, IFITM2, and IFITM3 in the inhibition of TBEV infection and in protection against virus-induced cell death. We show that the most significant role is that of IFITM3, including the dissection of its functional motifs by mutagenesis. Furthermore, through the use of CRISPR-Cas9-generated IFITM1/3-knockout monoclonal cell lines, we confirm the role and additive action of endogenous IFITMs in TBEV suppression. However, the results of coculture assays suggest that TBEV might partially escape interferon- and IFITM-mediated suppression during high-density coculture infection when the virus enters naive cells directly from infected donor cells. Thus, cell-to-cell spread may constitute a strategy for virus escape from innate host defenses. IMPORTANCE TBEV infection may result in encephalitis, chronic illness, or death. TBEV is endemic in northern Asia and Europe; however, due to climate change, new centers of endemicity have arisen. Although effective TBEV vaccines have been approved, vaccination coverage is low, and due to the lack of specific therapeutics, infected individuals depend on their immune responses to control the infection. IFITM proteins are components of the innate antiviral defenses that suppress cell entry of many viral pathogens. However, no studies on the role of IFITM proteins in TBEV infection have been published thus far. Understanding antiviral innate immune responses is crucial for the future development of antiviral strategies. Here, we show the important role of IFITM proteins in the inhibition of TBEV infection and virus-mediated cell death. However, our data suggest that TBEV cell-to-cell spread may be less prone to both interferon- and IFITM-mediated suppression, potentially facilitating escape from IFITM-mediated immunity.

Anti-HCV and Zika activities of ribavirin C-nucleosides analogues.

Ribavirin is an unnatural nucleoside exhibiting broad spectrum of antiviral and antitumor activities, still very widely studied particularly in a repositioning approach. C-triazolyl nucleoside analogues of ribavirin have been synthesized, as well as prodrugs and glycosylated or peptide conjugates to allow a better activity by vectorization into the liver or by facilitating uptake into the cells. The antiviral properties of all synthesized compounds have been evaluated in vitro against two important human viral pathogens belonging to the Flaviviridae family: hepatitis C virus (HCV) and Zika virus (ZIKV). There are no therapeutic options for Zika virus, whereas those available for HCV can be still improved. Our results indicated that compound 2 carrying an N-hydroxy carboxamide function exhibits the most inhibitory activities against both viruses. This compound moderately inhibited the propagation of HCV with an IC50 value of 49.1 µM and Zika virus with an IC50 of 33.2 µM comparable to ribavirin in the Vero cell line. The results suggest that compound 2 and its new derivatives may be candidates for further development of new anti-HCV and anti-ZIKV antiviral drugs.

Effects of Allyl Isothiocyanate on Oxidative and Inflammatory Stress in Type 2 Diabetic Rats.

Oxidative stress and inflammation play a crucial role in the pathogenesis and progression of diabetes. Currently, there is a growing need to exploit plant-derived bioactive compounds to support conventional therapies. The purpose of this study was to explore allyl isothiocyanate (AITC) potency in reducing oxidative and inflammatory stress along with its profitable modulation trace element status in pathological conditions such as diabetes. Two weeks of oral AITC treatments (2.5, 5, and 25 mg/kg body weight per day) were evaluated in Wistar rats with diabetes induced by a high-fat diet and streptozotocin. The study included AITC influence on antioxidant factors (SOD, CAT, GST, Nrf2), stress and inflammatory markers (cortisol, CRP, IL-1ß, IL-6, TNFα, NF-κB), lipid peroxidation indices (TBARS, -SH groups), and trace element status (Fe, Zn, and Cu) in the detoxification and lymphoid organs. Independently of dose, AITC increased cortisol levels in rat blood serum and decreased total thiol groups (T-SH) and protein-bound thiol groups (PB-SH) collaterally with raised thiobarbituric acid reactive substances (TBARS) in diabetic rat liver. The inflammation and oxidative effects were enhanced by an AITC dose increase. The highest dose of AITC, 25 mg/kg b.w., strongly affected the inflammation process by increasing IL-6, IL-1ß, and TNFα in the blood serum, and it upregulated Nrf2 transcription factor with increased SOD, GPx, and GST activities in the liver. AITC showed an equivocal effect on profitable modulation of disturbances in mineral homeostasis in the liver, kidney, and spleen. Our findings revealed that two-week AITC treatment exacerbated oxidative and inflammation status in diabetic rats.

Antiviral Activity of Uridine Derivatives of 2-Deoxy Sugars against Tick-Borne Encephalitis Virus.

Tick-borne encephalitis virus (TBEV) is a causative agent of tick-borne encephalitis (TBE), one of the most important human infections involving the central nervous system. Although effective vaccines are available on the market, they are recommended only in endemic areas. Despite many attempts, there are still no specific antiviral therapies for TBEV treatment. Previously, we synthesized a series of uridine derivatives of 2-deoxy sugars and proved that some compounds show antiviral activity against viruses from the Flaviviridae and Orthomyxoviridae families targeting the late steps of the N-glycosylation process, affecting the maturation of viral proteins. In this study, we evaluated a series of uridine derivatives of 2-deoxy sugars for their antiviral properties against two strains of the tick-borne encephalitis virus; the highly virulent TBEV strain Hypr and the less virulent strain Neudoerfl. Four compounds (2, 4, 10, and 11) showed significant anti-TBEV activity with IC50 values ranging from 1.4 to 10.2 µM and low cytotoxicity. The obtained results indicate that glycosylation inhibitors, which may interact with glycosylated membrane TBEV E and prM proteins, might be promising candidates for future antiviral therapies against TBEV.

Synthesis and Preliminary Evaluation of Biological Activity of Glycoconjugates Analogues of Acyclic Uridine Derivatives.

Herein we present the methodology for obtaining glycosyltransferase inhibitors, analogues of natural enzyme substrates of donor-type: UDP-glucose and UDP-galactose. The synthesis concerned glycoconjugates, nucleoside analogues containing an acyclic ribose mimetic linked to a uracil moiety in their structure. The biological activity of the synthesised compounds was determined on the basis of their ability to inhibit the model enzyme action of ß-1,4-galactosyltransferase from bovine milk. The obtained results allowed to expand and supplement the existing library of synthetic compounds that are able to regulate the biological activity of enzymes from the GT class.

Anti-Hepatitis C Virus Activity of Uridine Derivatives of 2-Deoxy Sugars.

Hepatitis C virus (HCV), the etiological agent of the most common and dangerous diseases of the liver, is a major health problem worldwide. Despite many attempts, there is still no vaccine available. Although many drugs have been approved for use mostly in combination regimen, their high costs make them out of reach in less developed regions. Previously, we have synthesized a series of compounds belonging to uridine derivatives of 2-deoxy sugars and have proved that some of them possess antiviral activity against influenza A virus associated with N-glycosylation inhibition. Here, we analyze the antiviral properties of these compounds against HCV. Using cell culture-derived HCV (HCVcc), HCV pseudoparticles (HCVpp), and replicon cell lines, we have shown high anti-HCV activity of two compounds. Our results indicated that compounds 2 and 4 significantly reduced HCVcc propagation with IC50 values in low µM range. Further experiments using the HCVpp system confirmed that both compounds significantly impaired the infectivity of produced HCVpp due to the inhibition of the correct maturation of viral glycoproteins. Overall, our results suggest that inhibiting the glycosylation process might be a good target for new therapeutics not only against HCV, but other important viral pathogens which contain envelopes with highly glycosylated proteins.

Novel Uridine Glycoconjugates, Derivatives of 4-Aminophenyl 1-Thioglycosides, as Potential Antiviral Compounds.

A novel series of uridine glycoconjugates, derivatives of 4-aminophenyl 1-thioglycosides, was designed and synthesized. All compounds were evaluated in vitro for their antiviral activity against hepatitis C virus (HCV) and classical swine fever virus (CSFV), two important human and animal viral pathogens for which new or improved therapeutic options are needed. The antiviral activity of all synthesized compounds was confirmed using pseudo-plaque reduction assays in which a significant arrest of CSFV and HCV growth was observed in the presence of these compounds. Two of the synthesized compounds, 9 and 12, displayed a significant inhibitory effect on HCV and CSFV propagation with IC50 values of 4.9 and 13.5 µM for HCV and 4.2 and 4 µM for CSFV, respectively, with low cytotoxicity. Using various infection and replication models, we have shown that both compounds were able to significantly reduce viral genome replication by up to 90% with IC50 values in the low micromolar range. A structure activity analysis of the synthesized compounds showed that the high antiviral activity was attributed to the hydrophobicity of glycoconjugates and the introduction of elements capable to coordinate metal ions into the spacer connecting the sugar and uridine moiety, which can be useful in the development of new antiviral compounds in the future.

Biological Evaluation of Uridine Derivatives of 2-Deoxy Sugars as Potential Antiviral Compounds against Influenza A Virus.

Influenza virus infection is a major cause of morbidity and mortality worldwide. Due to the limited ability of currently available treatments, there is an urgent need for new anti-influenza drugs with broad spectrum protection. We have previously shown that two 2-deoxy sugar derivatives of uridine (designated IW3 and IW7) targeting the glycan processing steps during maturation of viral glycoproteins show good anti-influenza virus activity and may be a promising alternative approach for the development of new anti-influenza therapy. In this study, a number of IW3 and IW7 analogues with different structural modifications in 2-deoxy sugar or uridine parts were synthesized and evaluated for their ability to inhibit influenza A virus infection in vitro. Using the cytopathic effect (CPE) inhibition assay and viral plaque reduction assay in vitro, we showed that compounds 2, 3, and 4 exerted the most inhibitory effect on influenza virus A/ostrich/Denmark/725/96 (H5N2) infection in Madin-Darby canine kidney (MDCK) cells, with 50% inhibitory concentrations (IC50) for virus growth ranging from 82 to 100 (µM) without significant toxicity for the cells. The most active compound (2) showed activity of 82 µM with a selectivity index value of 5.27 against type A (H5N2) virus. Additionally, compound 2 reduced the formation of HA glycoprotein in a dose-dependent manner. Moreover, an analysis of physicochemical properties of studied compounds demonstrated a significant linear correlation between lipophilicity and antiviral activity. Therefore, inhibition of influenza A virus infection by conjugates of uridine and 2-deoxy sugars is a new promising approach for the development of new derivatives with anti-influenza activities.

Effect of Elderberry (Sambucus nigra L.) Extract Supplementation in STZ-Induced Diabetic Rats Fed with a High-Fat Diet.

Elderberry (Sambucus nigra L.) lipophilic and polar extract dietary supplementation effects were evaluated according to diabetes management indices, using an in vivo model. A research pipeline was constructed, that ranged from extract preparation, partial chemical characterization and toxicity evaluation, to examining the elderberry extract dietary supplementation effects on biofluid and tissues. Extracts toxicity was screened using an Aliivibrio fischeri bioluminescence model. A concentration of up to 60 mg/L was selected, and rat doses for oral supplementation were computed applying the interspecies correlation between A. fischeri and rats. Wistar type 2 diabetic rats, induced by streptozotocin (STZ), were fed a high-fat diet and supplemented for 4 weeks at doses of 190 and 350 mg/kg body weight/day of lipophilic and polar extract, respectively. As far as we know, lipophilic elderberry extract supplementation was assessed for the first time, while polar extract was administrated at higher doses and for a shorter period compared to previous studies, aiming to evaluate subacute supplementation effects. The polar extract modulated glucose metabolism by correcting hyperglycemia, while the lipophilic extract lowered insulin secretion. Both extracts lowered insulin resistance, without remarkable alterations to hematological indices, sera lipids and sera and tissular trace element homeostasis. In conclusion, elderberries are a potential source of bioactive compounds for formulations to be used as co-adjuvants in diabetes management.

[Novel methods of hepatitis C treatment and prevention]. / Wirusowe zapalenie watroby typu C-nowe metody leczenia i zapobiegania.

Despite available treatment, Hepatitis C remains one of most serious burdens to public health. Current therapy based on pegylated interferon-alpha and ribavirin has significant side effects and its effectiveness varies for different genotypes of the virus. Four novel drugs - viral protease inhibitors (telaprevir, boceprevir, simeprevir) and polymerase inhibitor - sofosbuvir have been introduced in last years for use in combination with standard-of-care treatment. For the first time interferon free therapies were approved with the use of combination of sofosbuvir+ribavirin. New therapies improve virological response rates but also increase the cost, side effects and raise the issue of drug resistance. Numerous novel anti-HCV compounds have been evaluated in advanced clinical trials including inhibitors of viral proteins (protease, polymerase and NS5A) and inhibitors of host factors involved in HCV replication (cyclophilin A, microRNA - miR-122). New interferon-free therapies reducing severe side effects are expected to enter the market within few months. At the same time efforts are undertaken to determine the host and viral factors with predictive value for HCV treatment response, enabling personalized therapy approach. The main success in this field was the discovery of interleukin IL28B polymorphism, which correlates with positive standard-of-care treatment response. An effective vaccination may be an alternative for antiviral drugs, but no anti-HCV vaccine is available currently. It is well proved that successful vaccination should induce antibody and T-cell responses specific against a range of HCV genotypes. With this aim, new subunit and genetic candidate vaccines have been evaluated in I and II phase clinical trials. This review summarizes the recent developments in the field of new drug development and vaccine studies against hepatitis C virus.

Synthesis and antiviral activity of a novel glycosyl sulfoxide against classical swine fever virus.

A novel compound-2″,3″,4″,6″-tetra-O-acetyl-ß-d-galactopyranosyl-(1→4)-2',3',6'-tri-O-acetyl-1-thio-ß-d-glucopyranosyl-(5-nitro-2-pyridyl) sulfoxide-designated GP6 was synthesized and assayed for cytotoxicity and in vitro antiviral properties against classical swine fever virus (CSFV) in this study. We showed that the examined compound effectively arrested CSFV growth in swine kidney cells (SK6) at a 50% inhibitory concentration (IC50) of 5 ± 0.12 µg/ml without significant toxicity for mammalian cells. Moreover, GP6 reduced the viral E2 and E(rns) glycoproteins expression in a dose-dependent manner. We have excluded the possibility that the inhibitor acts at the replication step of virus life cycle as assessed by monitoring of RNA level in cells and culture medium of SK6 cells after single round of infection as a function of GP6 treatment. Using recombinant E(rns) and E2 proteins of classical swine fever virus produced in baculovirus expression system we have demonstrated that GP6 did not influence glycoprotein production and maturation in insect cells. In contrast to mammalian glycosylation pathway, insect cells support only the ER-dependent early steps of this process. Therefore, we concluded that the late steps of glycosylation process are probably the main targets of GP6. Due to the observed antiviral effect accompanied by low cytotoxicity, this inhibitor represents potential candidate for the development of antiviral agents for anti-flavivirus therapy. Further experiments are needed for investigating whether this compound can be used as a safe antiviral agent against other viruses from unrelated groups.

Influence of adjuvant type and route of administration on the immunogenicity of Leishmania-derived tick-borne encephalitis virus-like particles - A recombinant vaccine candidate.

Tick-borne encephalitis virus (TBEV) is a tick-borne flavivirus that induces severe central nervous system disorders. It has recently raised concerns due to an expanding geographical range and increasing infection rates. Existing vaccines, though effective, face low coverage rates in numerous TBEV endemic regions. Our previous work demonstrated the immunogenicity and full protection afforded by a TBEV vaccine based on virus-like particles (VLPs) produced in Leishmania tarentolae cells in immunization studies in a mouse model. In the present study, we explored the impact of adjuvants (AddaS03™, Alhydrogel®+MPLA) and administration routes (subcutaneous, intramuscular) on the immune response. Adjuvanted groups exhibited significantly enhanced antibody responses, higher avidity, and more balanced Th1/Th2 response. IFN-γ responses depended on the adjuvant type, while antibody levels were influenced by both adjuvant and administration routes. The combination of Leishmania-derived TBEV VLPs with Alhydrogel® and MPLA via intramuscular administration emerged as a highly promising prophylactic vaccine candidate, eliciting a robust, balanced immune response with substantial neutralization potential.

Influence of Dosing Regimen and Adjuvant Type on the Immunogenicity of Novel Recombinant Zika Virus-Like Particles.

Zika virus (ZIKV) is a reemerging mosquito-borne flavivirus that causes febrile illness and is also linked to Guillain-Barré syndrome as well as to microcephaly in newborns. Due to the risk of fetuses developing microcephaly, ZIKV is a serious problem for pregnant women. Although different types of vaccine antigens have been investigated, there is still no approved vaccine that prevents ZIKV. The aim of this study was to produce a potential anti-Zika virus vaccine candidate based on virus-like particles (VLPs) in mammalian cells and to analyze the role of dosing regimen and adjuvant type on the immunogenicity of the obtained antigen. Novel recombinant VLPs (F2A) were designed by introducing the optimized signal sequence of prM protein and by adding a self-cleavage peptide 2A between proteins prM and E. These modifications improved the formation of the glycoprotein E dimer. It has been shown that the increasing dosing regimen generates a significantly higher titer of antibodies; however, the adjuvant type does not affect this process. Sera from mice immunized using an increasing dosing schedule also showed higher neutralization activity against both Zika strains (H/PAN/2016/BEI-259634, a pandemic strain belonging to Asian lineage, and MR766, a reference strain from African lineage). In summary, this is the first report showing the influence of vaccination schedules and adjuvants on the immunogenicity of ZIKV virus-like particles. IMPORTANCE Considering the transmission of ZIKV and the risk of another epidemic as well as the neurological complications that follow ZIKV infection, the virus remains a serious problem for the human population, especially pregnant women. Therefore, there is a great need to develop new effective vaccine candidates. Although different types of vaccine antigens have been used in preclinical studies worldwide, there is still no approved vaccine to prevent ZIKV. VLPs are among the most potent antigens, but to use VLPs, adjuvants must be added to the formulation and appropriate administration must be performed. In this study, we show for the first time the influence of vaccination schedules and adjuvants on the immunogenicity of recombinant ZIKV VLPs. The obtained results can be used in new vaccine designs not only against ZIKV but also against other important viral pathogens.

Functional characterization and immunogenicity of a novel vaccine candidate against tick-borne encephalitis virus based on Leishmania-derived virus-like particles.

Tick-borne encephalitis virus (TBEV) is a major cause of neurological infections in many regions of central, eastern and northern Europe and northern Asia. In approximately 15% of cases, TBEV infections lead to the development of severe encephalitis or meningitis. The main route of TBEV transmission is tick bites; however, ingestion of dairy products from infected animals (goats, cattle and sheep) is also a frequent cause of the disease. Therefore, vaccination of livestock in virus endemic regions could also contribute to the decrease in TBEV infection among humans. Although few vaccines against TBEV based on inactivated viruses are available for humans, due to high costs, vaccination is not mandatory in most of the affected countries. Moreover, there is still no vaccine for veterinary use. Here, we present a characterization and immunogenicity study of a new potential TBEV vaccine based on virus-like particles (VLPs) produced in Leishmania tarentolae cells. VLPs, which mimic native viral particles but do not contain genetic material, show good immunogenic potential. For the first time, we showed that the protozoan L. tarentolae expression system can be successfully used for the production of TBEV virus-like particles with highly efficient production. We confirmed that TBEV recombinant structural proteins (prM/M and E) from VLPs are highly recognized by neutralizing antibodies in in vitro analyses. Therefore, VLPs in combination with AddaVax adjuvant were used in immunization studies in a mouse model. VLPs proved to be highly immunogenic and induced the production of high levels of neutralizing antibodies. In a challenge experiment, immunization with VLPs provided full protection from lethal TBE in mice. Thus, we suggest that Leishmania-derived VLPs may be a good candidate for a safe alternative human vaccine with high efficiency of production. Moreover, this potential vaccine candidate may constitute a low-cost candidate for veterinary use.

The Influence of Adjuvant Type on the Immunogenicity of RBD/N Cocktail Antigens as a Vaccine Candidate against SARS-CoV-2 Virus.

The emerging virus SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus 2 virus), agent of COVID-19, appeared in December 2019 in Wuhan, China, and became a serious threat to global health and public safety. Many COVID-19 vaccines have been approved and licensed around the world. Most of the developed vaccines include S protein and induce an antibody-based immune response. Additionally, T-cell response to the SARS-CoV-2 antigens could be beneficial for combating the infection. The type of immune response is greatly dependent not only on the antigen, but also on adjuvants used in vaccine formulation. Here, we compared the effect of four different adjuvants (AddaS03, Alhydrogel/MPLA, Alhydrogel/ODN2395, Quil A) on the immunogenicity of a mixture of recombinant RBD and N SARS-CoV-2 proteins. We have analyzed the antibody and T-cell response specific to RBD and N proteins and assessed the impact of adjuvants on virus neutralization. Our results clearly indicated that Alhydrogel/MPLA and Alhydrogel/ODN2395 adjuvants elicited the higher titers of specific and cross-reactive antibodies to S protein variants from various SARS-CoV-2 and SARS-CoV-1 strains. Moreover, Alhydrogel/ODN2395 stimulated high cellular response to both antigens, as assessed by IFN-γ production. Importantly, sera collected from mice immunized with RBD/N cocktail in combination with these adjuvants exhibited neutralizing activity against the authentic SARS-CoV-2 virus as well as particles pseudotyped with S protein from various virus variants. The results from our study demonstrate the immunogenic potential of RBD and N antigens and point out the importance of adjuvants selection in vaccine formulation in order to enhance the immunological response. IMPORTANCE Although several COVID-19 vaccines have been approved worldwide, continuous emergence of new SARS-CoV-2 variants calls for new efficient vaccines against them, providing long-lasting immunity. As the immune response after vaccination is dependent not only on antigen used, but also on other vaccine components, e.g., adjuvants, the purpose of this work was to study the effect of different adjuvants on the immunogenicity of RBD/N SARS-CoV-2 cocktail proteins. In this work, it has been shown that immunization with both antigens plus the different adjuvants studied elicited higher Th1 and Th2 responses against RBD and N, which contributed to higher neutralization of the virus. The obtained results can be used for design of new vaccines, not only against SARS-CoV-2, but also against other important viral pathogens.

Differential RNA editing landscapes in host cell versus the SARS-CoV-2 genome.

The SARS-CoV-2 pandemic was defined by the emergence of new variants formed through virus mutation originating from random errors not corrected by viral proofreading and/or the host antiviral response introducing mutations into the viral genome. While sequencing information hints at cellular RNA editing pathways playing a role in viral evolution, here, we use an in vitro human cell infection model to assess RNA mutation types in two SARS-CoV-2 strains representing the original and the alpha variants. The variants showed both different cellular responses and mutation patterns with alpha showing higher mutation frequency with most substitutions observed being C-U, indicating an important role for apolipoprotein B mRNA editing catalytic polypeptide-like editing. Knockdown of select APOBEC3s through RNAi increased virus production in the original virus, but not in alpha. Overall, these data suggest a deaminase-independent anti-viral function of APOBECs in SARS-CoV-2 while the C-U editing itself might function to enhance genetic diversity enabling evolutionary adaptation.

Oseltamivir analog with boron cluster modulator.

Synthesis of novel neuraminidase inhibitor -- carborane ester of oseltamivir carboxylic acid is described, and its physicochemical and spectral characteristics is provided. Surprisingly, carborane analog of oseltamivir is of an order of magnitude less active than its precursor, the corresponding ethyl ester, which is the active principle of pharmaceutical preparations used in influenza prophylactics and therapy.

Steviol Glycoside, L-Arginine, and Chromium(III) Supplementation Attenuates Abnormalities in Glucose Metabolism in Streptozotocin-Induced Mildly Diabetic Rats Fed a High-Fat Diet.

Stevia rebaudiana Bertoni and its glycosides are believed to exhibit several health-promoting properties. Recently, the mechanisms of the anti-diabetic effects of steviol glycosides (SG) have been the subject of intense research. The following study aims to evaluate the results of SG (stevioside (ST) and rebaudioside A (RA)) combined with L-arginine (L-Arg) and chromium(III) (CrIII) supplementation in streptozotocin- (STZ) induced mild type 2 diabetic rats fed a high-fat diet (HFD), with particular emphasis on carbohydrate and lipid metabolisms. The experiment was carried out on 110 male Wistar rats, 100 of which were fed an HFD to induce insulin resistance, followed by an intraperitoneal injection of streptozotocin to induce mild type 2 diabetes. After confirmation of hyperglycemia, the rats were divided into groups. Three groups served as controls: diabetic untreated, diabetic treated with metformin (300 mg/kg BW), and healthy group. Eight groups were fed an HFD enriched with stevioside or rebaudioside A (2500 mg/kg BW) combined with L-arginine (2000 or 4000 mg/kg BW) and Cr(III) (1 or 5 mg/kg BW) for six weeks. The results showed that supplementation with SG (ST and RA) combined with L-arg and Cr(III) could improve blood glucose levels in rats with mild type 2 diabetes. Furthermore, ST was more effective in improving blood glucose levels, insulin resistance indices, and very low-density lipoprotein cholesterol (VLDL-C) concentrations than RA. Although L-arg and Cr(III) supplementation did not independently affect most blood carbohydrate and lipid indices, it further improved some biomarkers when combined, particularly with ST. Notably, the beneficial impact of ST on the homeostatic model assessment-insulin resistance (HOMA-IR) and on the quantitative insulin-sensitivity check index (QUICKI) was strengthened when mixed with a high dose of L-arg, while its impact on antioxidant status was improved when combined with a high dose of Cr(III) in rats with mild type 2 diabetes. In conclusion, these results suggest that supplementary stevioside combined with L-arginine and Cr(III) has therapeutic potential for mild type 2 diabetes. However, further studies are warranted to confirm these effects in other experimental models and humans.

Effects of Bitter Melon and a Chromium Propionate Complex on Symptoms of Insulin Resistance and Type 2 Diabetes in Rat Models.

Trivalent chromium (Cr) and bitter melon (Momordica charantia L., BM) have been shown to independently interact with the insulin signaling pathway leading to improvements in the symptoms of insulin resistance and diabetes in some animal models and human subjects. The aim of this study was to examine whether the combination of the two nutritional supplements could potentially have additive effects on treating these conditions in high-fat-fed streptozotocin (STZ)-induced diabetic rats. The experiment was conducted with 110 male Wistar rats divided into eleven groups and fed either a control or high-fat diet for 7 weeks. Half of the rats on the high-fat diet were injected with STZ (30 mg/kg body mass) to induce diabetes. The high-fat (HF) diets were then supplemented with a combination of Cr (as chromium(III) propionate complex, Cr3: either 10 or 50 mg Cr/kg diet) and bitter melon (lyophilized whole fruit: either 10 or 50 g/kg diet) for 6 weeks. After termination of the experiment, blood and internal organs were harvested for blood biochemical, hematological, and mineral (Cr) analyses using appropriate analytical methods. It was found that neither Cr(III) nor BM was able to significantly affect blood indices in HF and diabetic rats, but BM tended to improve body mass gain, blood glucose, and LDL cholesterol values, but decreased Cr content in the liver and kidneys of the Cr-co-supplemented type 2 diabetic model of rats. Supplementary Cr(III) had no appreciable effect on glucose and lipid metabolism in high-fat-fed STZ-induced diabetic rats. Supplementary BM fruit powder had some observable effects on body mass of high-fat-fed rats; these effects seem to be dampened when BM was co-administered with Cr. Cr(III) and BM appear to act as nutritional antagonists when both administered in food, probably due to binding of Cr by the polyphenol-type compounds present in the plant material. Graphical Abstract.