An oral SARS-CoV-2 Mpro inhibitor clinical candidate for the treatment of COVID-19.

The worldwide outbreak of COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a global pandemic. Alongside vaccines, antiviral therapeutics are an important part of the healthcare response to countering the ongoing threat presented by COVID-19. Here, we report the discovery and characterization of PF-07321332, an orally bioavailable SARS-CoV-2 main protease inhibitor with in vitro pan-human coronavirus antiviral activity and excellent off-target selectivity and in vivo safety profiles. PF-07321332 has demonstrated oral activity in a mouse-adapted SARS-CoV-2 model and has achieved oral plasma concentrations exceeding the in vitro antiviral cell potency in a phase 1 clinical trial in healthy human participants.

Hard-to-heal wounds: a randomised trial of an oral proline-containing supplement to aid repair.

OBJECTIVE: We evaluated the effects of a specialised oral nutritional supplement (ONS) containing arginine and proline, with high vitamin A, C and E, zinc and selenium content, on the repair of hard-to-heal wounds. METHOD: Patients with hard-to-heal wounds were evaluated at five timepoints (S0-S4) over four consecutive weeks. At S0 patients were randomised to the specialised ONS (n=15; 25 wounds) or control (n=15; 25 wounds) groups. Posology was 200ml twice daily over the research period. Wound surface area and perimeter were monitored. In addition to the metric data, it was also possible to calculate the rate of wound contraction and the linear growth of the wound edges, looking for wound-healing predictive factors. RESULTS: A total of 30 patients took part in the study. Mean age was 65 years and 50% of patients had diabetes. Of the total evaluated wounds, 78% were <50cm2, 14% were 50-150cm2 and 8% were >250cm2. In 96% of cases, the wounds were in the lower limbs. A statistically significant reduction (p=0.004) in surface area of the wounds due to the specialised ONS, with a performance peak between S1 and S2, was observed. This specialised ONS did not induce changes in blood pressure, blood glucose level or renal function. A mean weekly wound edge growth of 1.85mm in patients with diabetes and 3.0mm in those without diabetes was observed. These results were 2.9 and 4.6 times, respectively, higher than expected, according to the literature. CONCLUSION: Specialised ONS can be a therapeutic option for hard-to-heal wounds.

Influence of Dietary Supplementation for Hyperhomocysteinemia Treatments.

Hyperhomocysteinemia is recognized as risk factor for cardiovascular and age-associated diseases. Folic acid supplementation efficiently lowers plasma homocysteine (Hcy) levels, but high intake may negatively affect health because of unnatural levels of unmetabolized folic acid in the systemic circulation. Oxoproline (Oxo) provides by glutamic acid production an increase of intracellular folic acid trapping. Aim of this study was to compare the efficacy of three supplementation protocols: (1) traditional therapy (5-methyl-tetrahydrofolate: 15 mg/day); (2) 5 mL/day of Oxo with 300 µg folic acid (oxifolic); (3) 5 mL/day of Oxo alone (magnesio+) in a 90 days randomized trial on thirty-two moderate hyperhomocysteinemic (18.6 ± 2.4 µmol.L-1) patients (age 48 ± 14 yrs). Thiols: cysteine (Cys), cysteinylglycine (Cys-Gly) and glutathione levels were assessed too. Every supplementation induced significant (p range <0.05-0.0001) reductions of Hcy level and Cys concentration after the three protocols adopted. Otherwise glutathione concentration significantly increased after oxifolic (p < 0.01) and traditional (p < 0.05) supplementation. The integration of Oxo resulted an interesting alternative to traditional therapy because absence or minimal number of folates in the integrator eliminates any chance of excess that can constitute a long-term risk.

Dietary supplementation with N-carbamylglycinate (CGly) improved feed source proline absorption and reproductive performance in sows.

Nutrient requirements are increased in the late-gestation period due to the faster growth of the foetal-placental unit and maternal erythrocyte mass. Glycine, proline and arginine are important amino acids that could improve foetal growth and development. The present study aims to investigate the effects of a derivative of glycine (N-carbamylglycinate, CGly) on the amino acid profiles and reproductive performances of late gestation sows. Thirty-two multiparous gestating sows (∼d 80) were selected, and randomly assigned into two groups: (1) control and (2) treatment (CGly, 800 mg kg-1) from day 85 of gestation to parturition. The serum amino acid profiles at day 110 of gestation and reproductive performance were investigated. The results showed that dietary supplementation of CGly in the late gestation period significantly improved the levels of glycine (p < 0.05) and proline (p < 0.01) in the serum of the perinatal sows, and thereafter improved the litter birth weights (p < 0.05) and number born alive (p < 0.1). Based on the in vitro studies, the improvement of proline levels is probably due to the induced expression of SLC6A20 and SLC38A2. Further studies should focus on the details of amino acid absorption, especially the competitive and cooperative absorption processes for different amino acids and derivatives.

Effects of maternal L-proline supplementation on inflammatory cytokines at the placenta and fetus interface of mice.

Dietary L-proline (proline) supplementation during gestation enhances fetal survival and placental development in mice. The objective of the present study was to test the hypothesis that this beneficial effect of proline was associated with alterations in inflammatory response at the placenta and fetus interface. Populations of immune cells present in peripheral blood mononuclear cells (PBMC) were determined by flow cytometry analysis. The concentrations of immunoglobulins in plasma, and the concentrations of cytokines in plasma, uterus, placenta, and amniotic fluid were measured using a bead-based immunoassay. The data showed that proline supplementation led to higher (P < 0.05) populations of B lymphocytes (CD3-CD19+), natural killer (NK) cells (CD3-NK1.1+), and dendritic cells (DCs, CD11c+MHCII+) in peripheral blood, as compared with the controls. Conversely, mice fed a proline-supplemented diet had a lower population of neutrophils (CD11b+F4/80-). Further study showed that proline supplementation decreased (P < 0.05) the concentrations of (1) interleukin (IL)-23, IL-1α, and IL-6 in plasma; (2) IL-6 in the uterus; and (3) tumor necrosis factor alpha (TNF-α), monocyte chemotactic protein (MCP)-1, and IL-17 in the placenta; and (4) interferon (IFN)-γ in amniotic fluid, compared with controls. Conversely, proline supplementation resulted in higher (P < 0.05) concentrations of (1) IL-10, IL-17 and granulocyte-macrophage colony-stimulating factor (GM-CSF) in plasma; (2) IL-10 and IL-1α in the uterus; and (3) IL-1α, IL-1ß, IL-10, IL-27, and IFN-ß in amniotic fluid, compared with controls. Moreover, concentrations of immunoglobulin (Ig) G2b and IgM were enhanced (P < 0.05) by proline administration. Taken together, our results reveal a regulatory effect of proline in the immunological response at the maternal-fetal interface, which is critical for embryonic development and fetal survival.

The Wound Healing Property of N-Methyl-(2S,4R)-trans-4-Hydroxy-L-Proline from Sideroxylon obtusifolium is Related to its Anti-Inflammatory and Antioxidant Actions.

Wound healing involves the interaction of blood cells, proteins, proteases, growth factors, and extracellular matrix components. Inflammation is one of the first events occurring during this process. Previously, we showed that the N-Methyl-(2S,4R)-trans-4-Hydroxy-L-Proline (NMP) from Sideroxylon obtusifolium leaves (a Brazilian medicinal species) presents an anti-inflammatory action. Considering inflammation as an important event in the wound healing process, the objectives were to investigate the topical effects of the NMP gel on a mice wound-induced model. Male Swiss mice were divided into 4 groups: Sham (surgical procedure only), Control (gel-base treated), and 3% or 10% NMP gel-treated groups. Measurements of wound areas and microscopic analyses (HE [hematoxylin-eosin] and PSR [picrosirius red] stainings) were carried out, at the 7th and 12th, days after the wound induction. Furthermore, immunohistochemical assays for iNOS (inducible nitric oxide synthase) and COX-2 (cyclooxygenase-2) and biochemical measurements for TBARS (thiobarbituric acid reactive substances), GSH (glutathione), and myeloperoxidase (MPO) were also performed, at the second day after the wound induction. The work showed that NMP decreases the wound areas, after topical application, relatively to the Sham and Control groups. In addition, microscopic alterations were reduced and collagen deposition was increased, at the 7th and 12th days, in the 10% NMP group. While iNOS and COX-2 immunostainings and GSH contents increased, in relation to the Sham and Control groups, TBARS and MPO decreased. Altogether, the results showed NMP to improve the wound healing process, by upregulating iNOS and COX-2 activities, reducing lipid peroxidation and MPO activity, and increasing GSH contents. In addition, NMP certainly contributes to the increased collagen deposition. These data may stimulate translational studies dealing with the possible use of NMP from Sideroxylon obtusifolium or from other sources for the management of wound healing.

Comparative pharmacokinetics study of leonurine and stachydrine in normal rats and rats with cold-stagnation and blood-stasis primary dysmenorrhoea after the administration of Leonurus japonicus houtt electuary.

Leonurus japonicus houtt, a well-known herb of traditional Chinese medicine, is widely used to treat gynaecological diseases. In this study, a rapid and sensitive liquid chromatography with tandem mass spectrometry method for simultaneously quantifying leonurine and stachydrine, the two main bioactive components in Leonurus japonicus houtt, was developed and validated. Plasma samples were prepared by protein precipitation with acetonitrile and separation by a Hewlett Packard XDB-C8 column (150 × 4.6 mm, id, 5 µm) equipped with a gradient elution system containing methanol-water and 0.1% formic acid at a flow-rate of 0.4 mL/min. Components were then detected by a mass spectrometer in positive electrospray ionization mode. This method showed good linearity, precision, accuracy, recovery, stability, and negligible matrix effects, which were within acceptable ranges. The method was successfully applied to compare the pharmacokinetics in normal rats and rats with cold-stagnation and blood-stasis primary dysmenorrhoea treated with Leonurus japonicus houtt electuary. The result showed significant differences (p < 0.05) in the pharmacokinetic parameters between the primary dysmenorrhoea and normal groups. This result implied that Leonurus japonicus houtt electuary remained longer and was absorbed slower in rats with primary dysmenorrhoea and exhibited higher bioavailability and peak concentration.

Maternal L-proline supplementation during gestation alters amino acid and polyamine metabolism in the first generation female offspring of C57BL/6J mice.

We recently reported that dietary supplementation with L-proline (proline) during gestation improved embryonic survival in C57BL/6J mice. The objective of the present study was to test the hypothesis that the effect of maternal proline supplementation on embryonic survival can be carried forward to the first generation female offspring. In the F0 generation, pregnant dams were fed a purified diet supplemented with 0 (control) or 5 g proline/kg diet. The F1 female adult offsprings were bred to fertile males. Fetal survival at embryonic day (E)12.5 and reproductive outcomes at term birth were recorded. The concentrations of amino acids, ammonia, and urea in plasma and amniotic fluid, as well as concentrations of polyamines in placental tissues and amniotic fluid at E12.5 were determined. Results showed that the F1 generation female offspring from proline-supplemented dams had higher (P < 0.05) concentrations of glutamate and taurine in plasma; of putrescine and spermidine in placental tissues; and of glycine, taurine, and spermidine in amniotic fluid at E12.5, as compared with F1 generation female offsprings from dams without proline supplementation. Concentration of proline in the plasma of offspring mice from proline-supplemented dams were lower (P < 0.05), as compared with the control group. No differences in fetal survival, reproductive outcomes, or concentrations of ammonia and urea in plasma and amniotic fluid were observed between the two groups of F1 female offspring. Collectively, our results indicate that the benefits of maternal proline supplementation during gestation on improving embryonic survival and fetal growth in F0 females are not transmitted to their F1 generation females.

l-proline supplementation improves nitric oxide bioavailability and counteracts the blood pressure rise induced by angiotensin II in rats.

We evaluated whether l-proline (Pro) supplementation improves redox status and nitric oxide (NO) bioavailability and prevents or delays angiotensin II (AngII)-induced hypertension. Male Sprague-Dawley rats were distributed to four experimental groups: Pro + AngII (Pro-Ang), Pro + Saline (Pro-Sal), Vehicle + AngII (Veh-Ang) and Veh + Saline (Veh-Sal). Pro solution (2 g.kg-1·day-1) or water (vehicle) were orally administered, from day 0 to day 21. AngII (200 ng.kg-1.min-1) or saline were infused (s.c.) from day 7 to day 21. Systolic blood pressure (SBP) was measured by the tail-cuff method. From day 20-21, animals were kept on metabolic cages for 24h-urine collection. On day 21, urine and blood were collected for further quantification of redox status biomarkers, NO-related markers (urinary nitrates and nitrites, U-NOx; plasma asymmetric dimethylarginine, P-ADMA), metabolic and renal parameters. Pro prevented the AngII-induced SBP rise [mean (95% CI), Day 19: Pro-AngII, 137 (131; 143) vs. Veh-AngII, 157 (151; 163) mm Hg, P < 0.001]. Pro-AngII rats also had increased values of U-NOx, systemic and urinary total antioxidant status (TAS), urinary H2O2 and plasma urea, as well as reduced P-ADMA and unaltered urinary isoprostanes. Plasma Pro was inversely correlated with P-ADMA (r = -0.52, p = 0.0009) and positively correlated with urinary TAS (r = 0.55, p = 0.0005) which, in turn, was inversely correlated with P-ADMA (r = -0.56, p = 0.0004). Furthermore, urinary H2O2 values decreased across P-ADMA tertiles (p for linear trend = 0.023). These results suggest that Pro reduces P-ADMA levels and improves redox status, thereby increasing NO bioavailability and counteracting the AngII-induced SBP rise. H2O2 and TAS modulation by Pro may contribute to the reduced P-ADMA concentration.

Combined Supplementation of Pre-Exercise Carbohydrate, Alanine, and Proline and Continuous Intake of Green Tea Catechins Effectively Boost Endurance Performance in Mice.

Continuous intake of green tea catechins (GTC) increases fatty acid utilization as an energy source and improves endurance capacity. Conversely, the single pre-exercise intake of maltodextrin (MD) as a carbohydrate source and the gluconeogenic amino acids alanine (Ala) and proline (Pro) effectively maintain blood glucose levels and increase endurance performance. In this study, we investigated the synergistic combinational effect of these interventions on endurance performance in mice. Male BALB/c mice were fed a 0.5% GTC diet or Control diet for 8 weeks. Maximum running time was measured every 2 weeks. MD (2 g/kg body weight (B.W.)), MD (1 g/kg B.W.) + AlaPro (9:1, 1 g/kg B.W.), and vehicle were orally administrated 60 mins before measurements in each diet group. The GTC + MD + AlaPro group showed significantly higher endurance performance than the Control-Vehicle group at all measurements. Indirect calorimetry analysis during running exercise at 4 weeks in the Control and GTC groups supplemented with pre-exercise MD + AlaPro administration revealed significantly higher fat oxidation in the GTC groups compared to the Control group. The combined increase in fatty acid utilization through continuous GTC intake and pre-exercise MD + AlaPro carbohydrate energy supplementation synergistically improves endurance capacity.

Maternal age modulates the effects of early-pregnancy L-proline supplementation on the birth-weight of piglets.

Previous results obtained in gilts maintained under experimental conditions suggest that amino acid supplementation during pregnancy may be a promising strategy for diminishing the incidence of embryo losses and low birth-weight newborn. The current study evaluated the effects of a short-term supplementation with L-proline, around implantational stages, on litter size and birth-weight of piglets in sows of different parities maintained under commercial farm conditions. There were no significant effects in mature sows with three or more parities, but the supplementation improved the reproductive efficiency of the high-prolific first-parity sows and of all the sows at second-parity. There were numerically higher litter size (of around two more live piglets; n.s.) and higher birth-weights (P<0.05) in the supplemented animals. The results of this study indicate that the effects of L-proline supplementation on litter size and birth-weight are strongly modulated by the maternal characteristics; specifically by parity and prolificacy and that supplementation may be cost-efficient for the management of females with compromised energy balance; specifically, sows at second farrowing and highly-prolific primiparous gilts.

Central mechanism of the cardiovascular responses caused by L-proline microinjected into the paraventricular nucleus of the hypothalamus in unanesthetized rats.

Previously, we reported that microinjection of L-proline (L-Pro) into the paraventricular nucleus of the hypothalamus (PVN) caused vasopressin-mediated pressor responses in unanesthetized rats. In the present study, we report on the central mechanisms involved in the mediation of the cardiovascular effects caused by the microinjection of L-Pro into the PVN. Microinjection of increasing doses of L-Pro (3-100nmol/100nL) into the PVN caused dose-related pressor and bradycardic responses. No cardiovascular responses were observed after the microinjection of equimolar doses (33nmol/100nL) of its isomer D-Proline (D-Pro) or Mannitol. The PVN pretreatment with either a selective non-NMDA (NBQX) or selective NMDA (LY235959 or DL-AP7) glutamate receptor antagonists blocked the cardiovascular response to L-Pro (33nmol/100nL). The dose-effect curve for the pretreatment with increasing doses of LY235959 was located at the left in relation to the curves for NBQX and DL-AP7, showing that LY235959 is more potent than NBQX, which is more potent than DL-AP7 in inhibiting the cardiovascular response to L-Pro. The cardiovascular response to the microinjection of L-Pro into the PVN was not affected by local pretreatment with Nω-Propyl-l-arginine (N-Propyl), a selective inhibitor of the neuronal nitric oxide synthase (nNOS), suggesting that NO does not mediate the responses to L-Pro in the PVN. In conclusion, the results suggest that ionotropic receptors in the PVN, blocked by both NMDA and non-NMDA receptor antagonists, mediate the pressor response to L-Pro that results from activation of PVN vasopressinergic magnocellular neurons and vasopressin release into the systemic circulation.

Efficacy and Safety of Topical Pidobenzone 4% as Adjuvant Treatment for Solar Lentigines: Result of a Randomized, Controlled, Clinical Trial.

BACKGROUND/OBJECTIVE: This study aims at the evaluation of the efficacy and safety of a combination therapy based on pidobenzone 4% and fractional CO2 laser or cryotherapy in the treatment of solar lentigines and the prevention of eventual posttreatment hyperchromia. METHODS: Efficacy was clinically evaluated by grading the pigmentation level with the Skin Tone Color Scale (STCS), and by grading patients' impression through a Visual Analog Scale (VAS). RESULTS: Our study shows that the associated treatment was safe and that it improves the therapeutic results on solar lentigines and prevents postiatrogenic hyperpigmentation compared with physical therapy alone. CONCLUSION: The combination of cryotherapy and pidobenzone 4% has been found to be the most useful treatment.

Maternally Administered Cyclic Glycine-Proline Increases Insulin-Like Growth Factor-1 Bioavailability and Novelty Recognition in Developing Offspring.

Cyclic glycine-proline (cGP), a metabolite of IGF-1, is an endogenous neuropeptide that improves memory in adult rats. The presence and concentrations of endogenous cGP, and its association with IGF-1 and IGF binding protein-3 (IGFBP-3) in rat milk and plasma, were evaluated during postnatal development. Maternal-infantile transfer of cGP during lactation and its efficacy on the memory of developing offspring were also investigated. Dams were gavaged with either cGP (3 mg/kg) or saline daily from postnatal days 8-22. Concentrations of cGP were measured in dams' milk, and concentrations of cGP, IGF-1, and IGFBP-3 were measured in the plasma of dams, pups, and young adults. The recognition memory, locomotor function, and anxiety-like behavior of offspring were evaluated using behavioral tests. Endogenous cGP was detected in rat milk, and its concentration was higher during peak lactation compared with late lactation. Comparisons within control groups showed low endogenous IGF-1 and IGFBP-3 and high endogenous cGP concentrations in the plasma of male pups. The reduced IGFBP-3 and increased cGP may be a response to increase the bioavailability of IGF-1 during infancy. Exogenous cGP showed oral bioavailability and effective maternal-infantile transfer through milk. Maternally transferred cGP also led to improved recognition memory in the developing offspring, possibly through increased IGF-1 bioavailability, with no effect on locomotor activity and anxiety-like behavior. These results show that cGP is an essential endogenous peptide during early postnatal development as it improves the bioavailability of IGF-1 during infancy. Furthermore, maternal cGP supplementation offers an effective and natural route of administration for improving memory in the developing offspring.

Polyphenols Inhibit Hepatitis C Virus Entry by a New Mechanism of Action.

UNLABELLED: Despite the validation of direct-acting antivirals for hepatitis C treatment, the discovery of new compounds with different modes of action may still be of importance for the treatment of special patient populations. We recently identified a natural molecule, epigallocatechin-3-gallate (EGCG), as an inhibitor of hepatitis C virus (HCV) targeting the viral particle. The aim of this work was to discover new natural compounds with higher anti-HCV activity than that of EGCG and determine their mode of action. Eight natural molecules with structure similarity to EGCG were selected. HCV JFH1 in cell culture and HCV pseudoparticle systems were used to determine the antiviral activity and mechanism of action of the compounds. We identified delphinidin, a polyphenol belonging to the anthocyanidin family, as a new inhibitor of HCV entry. Delphinidin inhibits HCV entry in a pangenotypic manner by acting directly on the viral particle and impairing its attachment to the cell surface. Importantly, it is also active against HCV in primary human hepatocytes, with no apparent cytotoxicity and in combination with interferon and boceprevir in cell culture. Different approaches showed that neither aggregation nor destruction of the particle occurred. Cryo-transmission electron microscopy observations of HCV pseudoparticles treated with delphinidin or EGCG showed a bulge on particles that was not observed under control conditions. In conclusion, EGCG and delphinidin inhibit HCV entry by a new mechanism, i.e., alteration of the viral particle structure that impairs its attachment to the cell surface. IMPORTANCE: In this article, we identify a new inhibitor of hepatitis C virus (HCV) infection, delphinidin, that prevents HCV entry. This natural compound, a plant pigment responsible for the blue-purple color of flowers and berries, belongs to the flavonoid family, like the catechin EGCG, the major component present in green tea extract, which is also an inhibitor of HCV entry. We studied the mode of action of these two compounds against HCV and demonstrated that they both act directly on the virus, inducing a bulging of the viral envelope. This deformation might be responsible for the observed inhibition of virus attachment to the cell surface. The discovery of such HCV inhibitors with an unusual mode of action is important to better characterize the mechanism of HCV entry into hepatocytes and to help develop a new class of HCV entry inhibitors.

L-proline increases survival of tilapias infected by Streptococcus agalactiae in higher water temperature.

Streptococcosis causes massive tilapia kills, which results in heavy economic losses of tilapia farming industry. Out of the Streptococcosis, Streptococcus agalactiae is the major pathogen. The bacterium causes higher mortality of tilapias in higher than lower temperatures. However, effect of temperature on metabolic regulation which is related to the mortality is largely unknown. The present study showed 50% and 70% mortality of tilapias cultured in 25 °C and 30 °C, respectively, in comparison with no death in 20 °C following infection caused by S. agalactiae. Then, GC/MS based metabolomics was used to investigate a global metabolic response of tilapia liver to the two higher water temperatures compared to 20 °C. Thirty-six and forty-five varied abundance of metabolites were identified in livers of tilapias cultured at 25 °C and 30 °C, respectively. More decreasing abundance of amino acids and increasing abundance of carbohydrates were detected in 30 °C than 25 °C groups. On the other hand, out of the pathways enriched, the first five biggest impact pathways belong to amino acid metabolism. Decreasing abundance of l-proline was identified as a crucial biomarker for indexing higher water temperature and a potential modulator to reduce the high death. This was validated by engineering injection or oral addition of l-proline. Exogenous l-proline led to elevated amino acid metabolism, which contributes to the elevated survivals. Our findings provide a potential metabolic modulator for controlling the disease, and shed some light on host metabolic prevention to infectious diseases.

In vitro inhibition of matrix metalloproteinases, invasion and growth of Fanconi anemia human FANCA and FANCC lymphoblasts by nutrient mixture.

AIM: Fanconi anemia is a rare genetic disorder with high propensity for development of cancers, such as aplastic anemia, leukemia and head and neck cancers. Collagen digesting matrix metalloproteinase (MMP) enzymes have been implicated in for their role in various malignancies and to promote metastasis. Biological agents that prevent extracellular matrix digestion by the MMPs have been shown to be promising therapeutic approaches to cancer. In this study, we investigated effects of a nutrient mixture (NM) containing, ascorbic acid, lysine, proline and green tea extract, on human FANCA and FANCC lymphoblasts for viability, MMP secretion and invasion. METHODS: Human FANCA lymphoblasts GM13022 and HCS536 were challenged with NM at concentration range within 10-1000 µg/ml. Cell toxicity was assessed by Trypan blue dye exclusion test. Invasion was evaluated through Matrigel and gelatinase zymography for MMP activity. RESULTS: NM was toxic in dose dependent mode to HCS536 cells but not to GM13022 cells. GM13022 cells but not HCS536 cells exhibited MMP-9 secretion, which was inhibited by NM. Matrigel invasion was inhibited in HCS536 cells at 100 and 500 µg/ml by 27% and 93%, respectively. In GM13022 cells, the NM showed completely blocked Matrigel invasion at 500 µg/ml. CONCLUSION: NM inhibited MMP secretion and Matrigel invasion in FANCA and inhibited invasion and induced toxicity in FANCC lymphoblasts. These results suggest that the NM may have therapeutic potential in Fanconi anemia associated neoplasia.

Combined supplementation of carbohydrate, alanine, and proline is effective in maintaining blood glucose and increasing endurance performance during long-term exercise in mice.

Carbohydrate supplementation is extremely important during prolonged exercise because it maintains blood glucose levels during later stages of exercise. In this study, we examined whether maintaining blood glucose levels by carbohydrate supplementation could be enhanced during long-term exercise by combining this supplementation with alanine and proline, which are gluconeogenic amino acids, and whether such a combination would affect exercise endurance performance. Male C57BL/6J mice were orally administered either maltodextrin (1.25 g/kg) or maltodextrin (1.0 g/kg) with alanine (0.225 g/kg) and proline (0.025 g/kg) 15 min before running for 170 min. Combined supplementation of maltodextrin, alanine, and proline induced higher blood glucose levels than isocaloric maltodextrin alone during the late exercise phase (100-170 min). The hepatic glycogen content of mice administered maltodextrin, alanine, and proline was higher than that of mice ingesting maltodextrin alone 60 min after beginning exercise, but the glycogen content of the gastrocnemius muscle showed no difference. We conducted a treadmill running test to determine the effect of alanine and proline on endurance performance. The test showed that running time to exhaustion of mice that were supplemented with maltodextrin (2.0 g/kg) was longer than that of mice that were supplemented with water alone. Maltodextrin supplementation (1.0 g/kg) with alanine (0.9 g/kg) and proline (0.1 g/kg) further increased running time to exhaustion compared to maltodextrin alone (2.0 g/kg). These results indicate that combined supplementation of carbohydrate, alanine, and proline is effective for maintaining blood glucose and hepatic glycogen levels and increasing endurance performance during long-term exercise in mice.

Meal pattern of male rats maintained on amino acid supplemented diets: the effect of tryptophan, lysine, arginine, proline and threonine.

The macronutrient composition of the diet has been shown to affect food intake, with proteins having distinct effects. The present study investigated the effect of diet supplementation with individual amino acids (tryptophan, lysine, arginine, proline and threonine) on meal pattern among male rats. Meal pattern and body weight were monitored for two weeks. Proline and threonine had minimal effects on meal pattern, while the most pronounced changes were observed in the tryptophan group. Both tryptophan and lysine decreased overall food intake, which was translated into a reduction in body weight. The reduced food intake of the tryptophan group was associated with an increase in meal size, intermeal intervals (IMI) and meal time and a decrease in meal number. The decrease in the food intake of the lysine group was associated with a reduction in both IMI and meal number, and this was accompanied by an increase in meal time. Arginine increased meal number, while decreasing IMI. Proline and threonine had a minimal effect on meal pattern. Lysine seems to increase satiety, and arginine seems to decrease it, while tryptophan seems to increase satiety and decrease satiation. Accordingly, changes in meal patterns are associated with the type of amino acid added to the diet.

Pharmacokinetics of the co-administration of boceprevir and St John's wort to male and female healthy volunteers.

BACKGROUND: St John's wort (SJW; Hypericum perforatum) induces CYP3A4 that is involved in the metabolism of the hepatitis C virus (HCV) protease inhibitor boceprevir. Reduced boceprevir exposure and efficacy would contribute to therapeutic failure and increase the risk for resistance development. Boceprevir is co-administered with interferon/ribavirin, and depression has been described frequently in patients undergoing HCV treatment. Patients may purchase over-the-counter herbals to manage depression, and knowing the interaction between SJW and boceprevir is desirable. METHODS: This Phase I, open-label, three-period, cross-over pharmacokinetic study enrolled healthy males and females who, following consent and screening procedures, were randomized to receive SJW on days 1-14, SJW plus boceprevir (SJW on days 22-35 and together on days 31-35) and boceprevir on days 52-56, separated by 7 day washout periods, or the same treatment in the opposite order. Pharmacokinetic sampling was performed at the end of each phase. RESULTS: Seventeen (11 female) subjects completed the study and no serious adverse events were reported. Geometric mean ratios (GMRs) and 90% CIs for boceprevir (with SJW versus alone) AUC(0-8), C(max) and C8 were 0.91 (0.87-0.96), 0.94 (0.82-1.07) and 1.00 (0.79-1.27), respectively. GMRs and 90% CIs for hypericin, the active component of SJW, (with boceprevir versus alone) AUC(0-8), C(max) and C(8) were 1.23 (1.10-1.38), 1.32 (1.16-1.52) and 1.37 (1.19-1.58), respectively. CONCLUSIONS: SJW did not have a clinically significant effect on boceprevir plasma concentrations (or those of its metabolite), suggesting that SJW and boceprevir can be safely co-administered.