Enhancing Curcumin's therapeutic potential in cancer treatment through ultrasound mediated liposomal delivery.

Improving the efficacy of chemotherapy remains a key challenge in cancer treatment, considering the low bioavailability, high cytotoxicity, and undesirable side effects of some clinical drugs. Targeted delivery and sustained release of therapeutic drugs to cancer cells can reduce the whole-body cytotoxicity of the agent and deliver a safe localized treatment to the patient. There is growing interest in herbal drugs, such as curcumin, which is highly noted as a promising anti-tumor drug, considering its wide range of bioactivities and therapeutic properties against various tumors. Conversely, the clinical efficacy of curcumin is limited because of poor oral bioavailability, low water solubility, instability in gastrointestinal fluids, and unsuitable pH stability. Drug-delivery colloid vehicles like liposomes and nanoparticles combined with microbubbles and ultrasound-mediated sustained release are currently being explored as effective delivery modes in such cases. This study aimed to synthesize and study the properties of curcumin liposomes (CLs) and optimize the high-frequency ultrasound release and uptake by a human breast cancer cell line (HCC 1954) through in vitro studies of culture viability and cytotoxicity. CLs were effectively prepared with particles sized at 81 ± 2 nm, demonstrating stability and controlled release of curcumin under ultrasound exposure. In vitro studies using HCC1954 cells, the combination of CLs, ultrasound, and Definity microbubbles significantly improved curcumin's anti-tumor effects, particularly under specific conditions: 15 s of continuous ultrasound at 0.12 W/cm2 power density with 0.6 × 107 microbubbles/mL. Furthermore, the study delved into curcumin liposomes' cytotoxic effects using an Annexin V/PI-based apoptosis assay. The treatment with CLs, particularly in conjunction with ultrasound and microbubbles, amplified cell apoptosis, mainly in the late apoptosis stage, which was attributed to heightened cellular uptake within cancer cells.

Rosemarinic acid protects ß-cell from STZ-induced cell damage via modulating NF-κß pathway.

Rosmarinic acid (RA), a natural ester phenolic compound, is known to have antioxidant and anti-inflammatory properties. RA has also been reported to exhibit a hypoglycemic effect; however, the mechanisms underlying this effect have yet to be investigated. Therefore, the present study focused on the anti-diabetic effects and mechanism of RA in INS-1 cells using in vitro model. Streptozotocin (STZ) at a concentration of 3 mM was applied to INS-1 cells for 4 h to create a diabetic model. The cells were pretreated for 24 h with various concentrations (1 and 2.5 µM) of RA. The Cell viability, glucose-stimulated insulin secretion (GSIS), glucose uptake, lipid peroxidation, reactive oxygen species (ROS), apoptosis, and protein expression of Bcl-2, NF-κB, 1L-1ß, and PARP were assessed. Results showed that STZ-treated INS-1 cells exhibited reduced cell viability, insulin release, insulin content, glucose uptake, and elevated MDA and ROS levels. Cells pretreated with RA maintained the function and morphology of ß-cells against STZ-induced damage. Moreover, RA sustained high protein expression levels of Bcl-2 and low expression levels of NF-κB, IL-1ß, and PARP. In conclusion, RA preserved ß-cells function against STZ-induced damage by altering NF-κB and Bcl-2 pathways.

Carnosic Acid Protects INS-1 ß-Cells against Streptozotocin-Induced Damage by Inhibiting Apoptosis and Improving Insulin Secretion and Glucose Uptake.

Carnosic acid (CA), a natural polyphenolic diterpene derived from Rosmarinus officinalis, has been proven to possess a broad spectrum of medicinal properties. Nevertheless, no studies on its impact on pancreatic ß-cells have been conducted to date. Herein, clonal rat INS-1 (832/13) cells were pretreated with CA for 24 h and then incubated with streptozotocin (STZ) for 3 h. Several functional experiments were performed to determine the effect of CA on STZ-induced pancreatic ß-cell damage, including cell viability assay, apoptosis analysis, and measurement of the level of insulin secretion, glucose uptake, malondialdehyde (MDA), reactive oxygen species (ROS), and proteins expression. STZ treatment decreased cell survival, insulin secretion, glucose uptake, and increased apoptosis, MDA, and ROS production in INS-1 cells. Furthermore, protein expression/phosphorylation analysis showed significant down-regulation in insulin, PDX-1, PI3K, AKT/p-AKT, and Bcl2. On the other hand, expression of BAX and BAD and cleaved PARP were significantly increased. Interestingly, preincubation with CA reversed the adverse impact of STZ at the cellular and protein expression levels. In conclusion, the data indicate that CA protects ß-cells against STZ-induced damage, presumably through its modulatory effect on the different pathways, including the Pi3K/AKT/PDX-1/insulin pathway and mitochondria-mediated apoptosis.

Central and peripheral neuropeptide RFRP-3: A bridge linking reproduction, nutrition, and stress response.

This article is an amalgamation of the current status of RFRP-3 (GnIH) in reproduction and its association with the nutrition and stress-mediated changes in the reproductive activities. GnIH has been demonstrated in the hypothalamus of all the vertebrates studied so far and is a well-known inhibitor of GnRH mediated reproduction. The RFRP-3 neurons interact with the other hypothalamic neurons and the hormonal signals from peripheral organs for coordinating the nutritional, stress, and environmental associated changes to regulate reproduction. RFRP-3 has also been shown to regulate puberty, reproductive cyclicity and senescence depending upon the nutritional status. A favourable nutritional status and the environmental cues which are permissive for the successful breeding and pregnancy outcome keep RFRP-3 level low, whereas unfavourable nutritional status and stressful conditions increase the expression of RFRP-3 which impairs the reproduction. Still our knowledge about RFRP-3 is incomplete regarding its therapeutic application for nutritional or stress-related reproductive disorders.

Copine 3 "CPNE3" is a novel regulator for insulin secretion and glucose uptake in pancreatic ß-cells.

Copine 3 (CPNE3) is a calcium-dependent phospholipid-binding protein that has been found to play an essential role in cancer progression and stages. However, its role in pancreatic ß-cell function has not been investigated. Therefore, we performed a serial of bioinformatics and functional experiments to explore the potential role of Cpne3 on insulin secretion and ß-cell function in human islets and INS-1 (832/13) cells. RNA sequencing and microarray data revealed that CPNE3 is highly expressed in human islets compared to other CPNE genes. In addition, expression of CPNE3 was inversely correlated with HbA1c and reduced in human islets from hyperglycemic donors. Silencing of Cpne3 in INS-1 cells impaired glucose-stimulated insulin secretion (GSIS), insulin content and glucose uptake efficiency without affecting cell viability or inducing apoptosis. Moreover, mRNA and protein expression of the key regulators in glucose sensing and insulin secretion (Insulin, GLUT2, NeuroD1, and INSR) were downregulated in Cpne3-silenced cells. Taken together, data from the present study provides a new understanding of the role of CPNE3 in maintaining normal ß-cell function, which might contribute to developing a novel target for future management of type 2 diabetes therapy.

The Coffee Diterpene, Kahweol, Ameliorates Pancreatic ß-Cell Function in Streptozotocin (STZ)-Treated Rat INS-1 Cells through NF-kB and p-AKT/Bcl-2 Pathways.

Kahweol is a diterpene molecule found in coffee that exhibits a wide range of biological activity, including anti-inflammatory and anticancer properties. However, the impact of kahweol on pancreatic ß-cells is not known. Herein, by using clonal rat INS-1 (832/13) cells, we performed several functional experiments including; cell viability, apoptosis analysis, insulin secretion and glucose uptake measurements, reactive oxygen species (ROS) production, as well as western blotting analysis to investigate the potential role of kahweol pre-treatment on damage induced by streptozotocin (STZ) treatment. INS-1 cells pre-incubated with different concentrations of kahweol (2.5 and 5 µM) for 24 h, then exposed to STZ (3 mmol/L) for 3 h reversed the STZ-induced effect on cell viability, apoptosis, insulin content, and secretion in addition to glucose uptake and ROS production. Furthermore, Western blot analysis showed that kahweol downregulated STZ-induced nuclear factor kappa B (NF-κB), and the antioxidant proteins, Heme Oxygenase-1 (HMOX-1), and Inhibitor of DNA binding and cell differentiation (Id) proteins (ID1, ID3) while upregulated protein expression of insulin (INS), p-AKT and B-cell lymphoma 2 (BCL-2). In conclusion, our study suggested that kahweol has anti-diabetic properties on pancreatic ß-cells by suppressing STZ induced apoptosis, increasing insulin secretion and glucose uptake. Targeting NF-κB, p-AKT, and BCL-2 in addition to antioxidant proteins ID1, ID3, and HMOX-1 are possible implicated mechanisms.

RF-amide related peptide-3 (RFRP-3): a novel neuroendocrine regulator of energy homeostasis, metabolism, and reproduction.

A hypothalamic neuropeptide, RF-amide related peptide-3 (RFRP-3), the mammalian ortholog of the avian gonadotropin-inhibitory hormone (GnIH) has inhibitory signals for reproductive axis via G-protein coupled receptor 147 in mammals. Moreover, RFRP-3 has orexigenic action but the mechanism involved in energy homeostasis and glucose metabolism is not yet known. Though, the RFRP-3 modulates orexigenic action in co-operation with other neuropeptides, which regulates metabolic cues in the hypothalamus. Administration of GnIH/RFRP-3 suppresses plasma luteinizing hormone, at the same time stimulates feeding behavior in birds and mammals. Likewise, in the metabolically deficient conditions, its expression is up-regulated suggests that RFRP-3 contributes to the integration of energy balance and reproduction. However, in many other metabolic conditions like induced diabetes and high-fat diet obesity, etc. its role is still not clear while, RFRP-3 induces the glucose homeostasis by adipocytes is reported. The physiological role of RFRP-3 in metabolic homeostasis and the metabolic effects of RFRP-3 signaling in pharmacological studies need a detailed discussion. Further studies are required to find out whether RFRP-3 is associated with restricted neuroendocrine function observed in type II diabetes mellitus, aging, or sub-fertility. In this context, the current review is focused on the role of RFRP-3 in the above-mentioned mechanisms. Studies from search engines including PubMed, Google Scholar, and science.gov are included after following set inclusion/exclusion criteria. As a developing field few mechanisms are still inconclusive, however, based on the available information RFRP-3 seems to be a putative tool in future treatment strategies towards metabolic disease.

Possible Role of GnIH as a Mediator between Adiposity and Impaired Testicular Function.

The aim of the present study was to evaluate the roles of gonadotropin-inhibitory hormone (GnIH) as an endocrine link between increasing adiposity and impaired testicular function in mice. To achieve this, the effect of GnIH on changes in nutrients uptake and hormonal synthesis/action in the adipose tissue and testis was investigated simultaneously by in vivo study and separately by in vitro study. Mice were treated in vivo with different doses of GnIH for 8 days. In the in vitro study, adipose tissue and testes of mice were cultured with different doses of GnIH with or without insulin or LH for 24 h at 37°C. The GnIH treatment in vivo showed increased food intake, upregulation of glucose transporter 4 (GLUT4), and increased uptake of triglycerides (TGs) in the adipose tissue. These changes may be responsible for increased accumulation of fat in white adipose tissue, resulting in increase in the body mass. Contrary to the adipose tissue, treatment with GnIH both in vivo and in vitro showed decreased uptake of glucose by downregulation of glucose transporter 8 (GLUT8) expressions in the testis, which in turn resulted in the decreased synthesis of testosterone. The GnIH treatment in vivo also showed the decreased expression of insulin receptor protein in the testis, which may also be responsible for the decreased testicular activity in the mice. These findings thus suggest that GnIH increases the uptake of glucose and TGs in the adipose tissue, resulting in increased accumulation of fat, whereas simultaneously in the testis, GnIH suppressed the GLUT8-mediated glucose uptake, which in turn may be responsible for decreased testosterone synthesis. This study thus demonstrates GnIH as mediator of increasing adiposity and impaired testicular function in mice.

Inhibitory roles of the mammalian GnIH ortholog RFRP3 in testicular activities in adult mice.

The aim of this study was to evaluate the effects of in vivo and in vitro treatments with RFamide-related peptide 3 (RFRP3), a mammalian gonadotropin-inhibitory hormone ortholog, on testicular activities, i.e. spermatogenesis and steroidogenesis, in mice. Mice were treated in vivo with different doses of RFRP3 (control: 0.02 µg, 0.2 µg, and 2.0 µg/day) for 8 days. For in vitro study, the testes of mice were evaluated with different doses of RFRP3 (control: 1 and 10 ng/ml) with or without LH (control: 10 and 100 ng/ml) for 24 h at 37 °C. RFRP3 treatment produced significant changes in the body mass, circulating steroid level, and testicular activity in mice. RFRP3 treatment also caused dose-dependent histological changes in spermatogenesis, such as decline in germ cell proliferation and survival markers and increase in apoptotic markers in testis. Both in vivo and in vitro studies showed the inhibitory effect of RFRP3 on testosterone synthesis in the testis. RFRP3 inhibited the expression of the receptor for LH (LHCGR), STAR protein, cytochrome P450 side-chain cleavage (CYP11A1) and 3ß-hydroxysteroid dehydrogenase in the testis, and testosterone secretion dose dependently. This study also suggested that the inhibitory effect of RFRP3 in the testis may be mediated through local production of GnRH. Thus, RFRP3 inhibits testicular steroidogenesis and spermatogenesis either indirectly through GnRH or by directly influencing germ cell proliferation, survival, and apoptosis.

Localization of gonadotropin-releasing hormone (GnRH), gonadotropin-inhibitory hormone (GnIH), kisspeptin and GnRH receptor and their possible roles in testicular activities from birth to senescence in mice.

The changes in distribution and concentration of neuropeptides, gonadotropin-releasing hormone (GnRH), gonadotropin-inhibitory hormone (GnIH), kisspeptin, and gonadotropin-releasing hormone receptor (GnRH-R) were evaluated and compared with reproductive parameters, such as cytochrome P450 side-chain cleavage (P450 SCC) enzyme activity, androgen receptors (AR) in the testis and serum testosterone levels, from birth to senescence in mice. The results showed the localization of these molecules mainly in the interstitial and germ cells as well as showed significant variations in immunostatining from birth to senescence. It was found that increased staining of testicular GnRH-R coincided with increased steroidogenic activity during pubertal and adult stages, whereas decreased staining coincides with decreased steroidogenic activity during senescence. Similar changes in immunostaining were confirmed by Western/slot blot analysis. Thus, these results suggest a putative role of GnRH during testicular pubertal development and senescence. Treatment with a GnRH agonist ([DTrp(6), Pro(9)-NEt] GnRH) to mice from prepubertal to pubertal period showed a significant increase in steroidogenic activity of the mouse testis and provided further support to the role of GnRH in testicular pubertal maturation. The significant decline in GnRH-R during senescence may be due to a significant increase in GnIH synthesis during senescence causing the decrease in GnRH-R expression. It is considered that significant changes in the levels of GnRH-R may be responsible for changes in steroidogenesis that causes either pubertal activation or senescence in testis of mice. Furthermore, changes in the levels of GnRH-R may be modulated by interactions among GnRH, GnIH, and kisspeptin in the testis.

Alteration in expression of estrogen receptor isoforms alpha and beta, and aromatase in the testis and its relation with changes in nitric oxide during aging in mice.

The aim of present study was to investigate the changes in the testicular expression of aromatase, ER alpha, ER beta and iNOS protein and correlate these with serum testosterone and nitric oxide levels, to elucidate the role of estrogen and nitric oxide in the testis during aging. This study showed localization of aromatase and ER alpha mainly in the Leydig cell and showed close correlation of testicular aromatase level with circulating testosterone level suggesting that estrogen may be modulating testicular steroidogenesis. Localization ER alpha mainly in the mitotically active germ cell suggest possible role of estrogen in germ cell proliferation. This study showed basal level of nitric oxide during reproductively active period, whereas increased serum nitric oxide coincides with decreased testicular activity in old age. This study showed inverse correlation between aromatase and NO level. Treatment with either SNP or L-NAME on testicular steroidogenic factor (3-beta HSD/ StAR) or germ cell survival factor (Bcl2) showed that increased NO causes decreased steroidogenesis and increased germ cell apoptosis. In conclusion this study suggest that estrogen modulate steroidogenesis and germ cell survival in reproductively active period whereas in old age decreased estrogen concentration causes increased nitric oxide which in turn decreases testicular steroidogenesis and germ cell apoptosis.