Safety of Oral Carica papaya L. Leaf 10% Ethanolic Extract for Acute and Chronic Toxicity Tests in Sprague Dawley Rats.

Carica papaya L. leaves, traditionally utilized in dietary supplements and pharmaceuticals, exhibit a broad spectrum of potentially therapeutic properties, including anti-inflammatory, antimalarial, and wound healing properties. This study examined the acute and chronic toxicity of 10% ethanolic-extracted C. papaya leaf in Sprague Dawley rats. The acute toxicity assessment was a single oral dose of 5000 mg/kg body weight, while the chronic toxicity assessment included daily oral doses of 100, 400, 1000, and 5000 mg/kg over 180 days. Systematic monitoring covered a range of physiological and behavioral parameters, including body and organ weights. End-point evaluations encompassed hematological and biochemical analyses, along with gross and histopathological examinations of internal organs. Findings revealed no acute toxicity in the C. papaya leaf extract group, although a significant decrease in uterine weight was observed without accompanying histopathology abnormalities. In the chronic toxicity assessment, no statistically significant differences between the control and the C. papaya leaf extract groups were detected across multiple measures, including behavioral, physiological, and hematological indices. Importantly, histopathological examination corroborated the absence of any tissue abnormalities. The study results indicate that C. papaya leaf extract exhibited no adverse effects on the rats during the 180-day oral administration period, affirming its potential safety for prolonged usage.

Clerodendrum chinense Stem Extract and Nanoparticles: Effects on Proliferation, Colony Formation, Apoptosis Induction, Cell Cycle Arrest, and Mitochondrial Membrane Potential in Human Breast Adenocarcinoma Breast Cancer Cells.

Breast cancer stands out as the most widespread form of cancer globally. In this study, the anticancer activities of Clerodendrum chinense (C. chinense) stem ethanolic extract were investigated. High-performance liquid chromatography (HPLC) analysis identified verbascoside and isoverbascoside as the major bioactive compounds in the C. chinense stem extract. Successfully developed nanoparticles exhibited favorable hydrodynamic diameter, polydispersity index, and surface charge, thus ensuring stability after four months of storage. The total phenolic content and total flavonoid contents in the nanoparticles were reported as 88.62% and 95.26%, respectively. The C. chinense stem extract demonstrated a dose-dependent inhibitory effect on MCF-7, HeLa, A549, and SKOV-3 cancer cell lines, with IC50 values of 109.2, 155.6, 206.9, and 423 µg/mL, respectively. C. chinense extract and NPs exhibited dose-dependent cytotoxicity and the highest selectivity index values against MCF-7 cells. A dose-dependent reduction in the colony formation of MCF-7 cells was observed following treatment with the extract and nanoparticles. The extract induced cytotoxicity in MCF-7 cells through apoptosis and necrosis. C. chinense stem extract and nanoparticles decreased mitochondrial membrane potential (MMP) and induced G0/G1 phase arrest in MCF-7 cells. In conclusion, use of C. chinense stem extract and nanoparticles may serve as a potential therapeutic approach for breast cancer, thus warranting further exploration.

The Identification and Cytotoxic Evaluation of Nutmeg (Myristica fragrans Houtt.) and Its Substituents.

The aril and seed of nutmeg, Myristica fragrans Houtt. (Myristicaceae), hold significant value in various industries globally. Our preliminary research found two morphological variations: a globose shape and an oval shape. Due to these different characteristics, the safety of consumers is of primary concern. Thus, authentication and comparative pharmacological and toxicity analyses are necessary. In this study, pharmacognostic and advanced phytochemical analyses, DNA barcoding, cytotoxicity, and the anti-nitric oxide production of commercial Thai nutmeg were examined. Via morphologic examinations and TLC fingerprinting, all the sampled aril and seed were categorized into globose and oval-shaped groups. The results of HPLC, GC-MS, and LC-MS/MS experiments revealed distinct differences between these groups. The DNA barcoding of the trnH-psbA region using the BLAST method and neighbor-joining tree analyses confirmed the globose nutmeg as M. fragrans and the oval-shaped variant as M. argentea. A comparison was then carried out between the potential toxicity and anti-inflammatory capabilities of M. fragrans and M. argentea. Cytotoxicity tests on HaCaT, 3T3-L1, Caco-2, HEK293, and RAW264.7 were performed using both methanolic extracts and volatile oil from the arils and seeds of both species. This study concludes that blending or substituting these two species maintains their therapeutic integrity without posing safety concerns.

Boesenbergia rotunda extract accelerates human keratinocyte proliferation through activating ERK1/2 and PI3K/Akt kinases.

Boesenbergia rotunda (BR) has long been used as tradition medicine. For its pharmacological effects on wound healing, previous studies in an animal model provided convincing results that the ethanolic extract from the rhizome of this plant can stimulate wound healing. However, the mechanism about how this plant promotes wound healing at the molecular level has not been elucidated. As a step towards the development of wound healing agents, our current study utilized a human keratinocyte cell line (HaCaT) as an in vitro model to define the potential molecular mechanisms of BR extract in enhancing wound-healing. Our HPLC results showed that BR extract contained kaempferol as one of its potential compounds. The extract strongly promoted wound healing of HaCaT cell monolayer. This effect was eventually defined to be regulated through the ability of BR extract to induce cell proliferation. At the signaling level, we discovered that BR extract rapidly activated ERK1/2 and Akt phosphorylation upon the addition of the extract. Additionally, our experiments where specific inhibitors of MEK (U0126) and PI3K (LY294002) were utilized verified that BR enhanced cell proliferation and wound healing through stimulating the MAPK and PI3K/Akt signal transduction pathways. Moreover, direct inhibition of keratinocyte DNA synthesis by mitomycin C (MMC) could completely block the proliferative effects of BR extract. Nevertheless, data from Transwell migration assay revealed that BR extract did not promote keratinocyte migration. Altogether, we provided more evidence that BR possesses its wound healing-promoting action through the activation of proliferation and survival pathways, and our study suggests that BR is an interesting candidate to be developed as a wound healing-promoting agent.

Artocarpus lakoocha Extract Inhibits LPS-Induced Inflammatory Response in RAW 264.7 Macrophage Cells.

:Artocarpus lakoocha Roxb. (AL) has been known for its high content of stilbenoids, especially oxyresveratrol. AL has been used in Thai traditional medicine for centuries. However, the role of AL in regulating inflammation has not been elucidated. Here we investigated the molecular mechanisms underlying the anti-inflammation of AL ethanolic extract in RAW 264.7 murine macrophage cell line. The HPLC results revealed that this plant was rich in oxyresveratrol, and AL ethanolic extract exhibited anti-inflammatory properties. In particular, AL extract decreased lipopolysaccharide (LPS)-mediated production and secretion of cytokines and chemokine, including IL-6, TNF-α, and MCP-1. Consistently, the extract inhibited the production of nitric oxide (NO) in the supernatants of LPS-stimulated cells. Data from the immunofluorescence study showed that AL extract suppressed nuclear translocation of nuclear factor-kappa B (NF-κB) upon LPS induction. Results from Western blot analysis further confirmed that AL extract strongly prevented the LPS-induced degradation of IκB which is normally required for the activation of NF-κB. The protein expression of iNOS and COX-2 in response to LPS stimulation was significantly decreased with the presence of AL extract. AL extract was found to play an anti-inflammatory role, in part through inhibiting LPS-induced activation of Akt. The extract had negligible impact on the activation of mitogen-activated protein kinase (MAPK) pathways. Specifically, incubation of cells with the extract for only 3 h demonstrated the rapid action of AL extract on inhibiting the phosphorylation of Akt, but not ERK1/2. Longer exposure (24 h) to AL extract was required to mildly reduce the phosphorylation of ERK1/2, p38, and JNK MAPKs. These results indicate that AL extract manipulates its anti-inflammatory effects mainly through blocking the PI3K/Akt and NF-κB signal transduction pathways. Collectively, we believe that AL could be a potential alternative agent for alleviating excessive inflammation in many inflammation-associated diseases.

Effect of Short-Course Oral Ciprofloxacin on Isoflavone Pharmacokinetics following Soy Milk Ingestion in Healthy Postmenopausal Women.

Soy isoflavones have several potential benefits related to postmenopausal health. Isoflavone glycosides, found predominantly in nonfermented soy products, e.g., soy milk, require conversion by gut microbiota to their respective bioavailable aglycones prior to absorption into portal circulation. Use of short-course oral ciprofloxacin for the treatment of acute uncomplicated cystitis, the incidence of which is increasing among postmenopausal women, might adversely affect gut microbiota. The objective of this one-group pre-post treatment study was to determine the effect of short-course oral ciprofloxacin on isoflavone pharmacokinetics in healthy postmenopausal women. Eleven postmenopausal subjects were assigned to consume a single oral dose of 375 mL UHT soy milk (SOY phase). Blood samples were collected immediately before soy milk ingestion and at specific times for 32 hours after soy milk ingestion. Following a washout period of at least seven days, subjects were assigned to take 250 mg oral ciprofloxacin after breakfast and dinner for three days, followed by a single oral dose of 375 mL UHT soy milk the next day (CIPRO/SOY phase). Blood samples were collected at the same time points as in the SOY phase. Plasma samples were treated with ß-glucuronidase/sulfatase and plasma concentrations of aglycones (genistein and daidzein) were determined using high-performance liquid chromatography. Cmax, AUC0-t, and AUC0-∞ of both aglycones and Tmax of genistein obtained from the CIPRO/SOY phase were significantly lower than those obtained from the SOY phase, while Tmax of daidzein and t1/2 of both aglycones in the two phases were not significantly different.

Anti-cancer effects of Kaempferia parviflora on ovarian cancer SKOV3 cells.

BACKGROUND: Kaempferia parviflora (KP) is an herb found in the north of Thailand and used as a folk medicine for improving vitality. Current reports have shown the anti-cancer activities of KP. However, the anti-cancer effects of KP on highly aggressive ovarian cancer have not been investigated. Therefore, we determined the effects of KP on cell proliferation, migration, and cell death in SKOV3 cells. METHODS: Ovarian cancer cell line, SKOV3 was used to investigate the anti-cancer effect of KP extract. Cell viability, cell proliferation, MMP activity, cell migration, and invasion were measured by MTT assay, cell counting, gelatin zymography, wound healing assay, and Transwell migration and invasion assays, respectively. Cell death was determined by trypan blue exclusion test, AnnexinV/PI with flow cytometry, and nuclear staining. The level of ERK and AKT phosphorylation, and caspase-3, caspase-7, caspase-9 was investigated by western blot analysis. RESULTS: KP extract was cytotoxic to SKOV3 cells when the concentration was increased, and this effect could still be observed even though EGF was present. Besides, the cell doubling time was significantly prolonged in the cells treated with KP. Moreover, KP strongly suppressed cell proliferation, cell migration and invasion. These consequences may be associated with the ability of KP in inhibiting the activity of MMP-2 and MMP-9 assayed by gelatin zymography. Moreover, KP at high concentrations could induce SKOV3 cell apoptosis demonstrated by AnnexinV/PI staining and flow cytometry. Consistently, nuclear labelling of cells treated with KP extract showed DNA fragmentation and deformity. The induction of caspase-3, caspase-7, and caspase-9 indicates that KP induces cell death through the intrinsic apoptotic pathway. The antitumor activities of KP might be regulated through PI3K/AKT and MAPK pathways since the phosphorylation of AKT and ERK1/2 was reduced. CONCLUSIONS: The inhibitory effects of KP in cell proliferation, cell migration and invasion together with apoptotic cell death induction in SKOV3 cells suggest that KP has a potential to be a new candidate for ovarian cancer chemotherapeutic agent.

Antipsoriatic Effects of Wannachawee Recipe on Imiquimod-Induced Psoriasis-Like Dermatitis in BALB/c Mice.

Psoriasis is a common immune-mediated chronic inflammatory skin disease characterized by thick and erythema raised plaques with adherent silvery scales. T-cells are activated via the IL-23/Th17 axis which is involved in psoriasis pathogenesis. Conventional treatments of psoriasis have adverse events that influence patients' adherence. Wannachawee Recipe (WCR) is Thai traditional medicine that is known to be effective for psoriasis patients; however, preclinical evidence is still lacking. This study investigated the therapeutic potential of WCR on antiproliferant activity using imiquimod- (IMQ-) induced psoriasis-like dermatitis in a mouse model. Psoriasis-like dermatitis was induced on the shaved dorsal skin and right ear pinna of BALB/c mice by topical application of IMQ for 15 consecutive days after which WCR was administered to the mice by oral gavage for 10 days. Phenotypical observations, histopathological examinations, and ELISA of skin and blood samples were conducted. WCR significantly ameliorated development of IMQ-induced psoriasis-like dermatitis and reduced levels of Th17 cytokines (IL-17A, IL-22, and IL-23) in both serum and dorsal skin. Histopathological findings showed a decrease in epidermal thickness and inflammatory T-cell infiltration in the WCR-treated groups. The WCR has pharmacological actions which regulate Th17 related cytokines suggesting that it is a potential alternative therapeutic strategy for psoriasis.

Effects of Wannachawee Recipe with Antipsoriatic Activity on Suppressing Inflammatory Cytokine Production in HaCaT Human Keratinocytes.

Psoriasis is a chronic inflammatory and immune-mediated skin disease. The pathogenesis involves T cells activation via the IL-23/Th17 axis. Conventional treatments of psoriasis have adverse events influencing patients' adherence. Wannachawee Recipe (WCR) has been effectively used as Thai folk remedy for psoriasis patients; however, preclinical evidence defining how WCR works is still lacking. This study defined mechanisms for its antiproliferation and anti-inflammatory effects in HaCaT cells. The cytotoxicity and antiproliferation results from SRB and CCK-8 assays showed that WCR inhibited the growth and viability of HaCaT cells in a concentration-dependent manner. The distribution of cell cycle phases determined by flow cytometry showed that WCR did not interrupt cell cycle progression. Interestingly, RT-qPCR revealed that WCR significantly decreased the mRNA expression of IL-1ß, IL-6, IL-8, IL-17A, IL-22, IL-23, and TNF-α but induced IL-10 expression in TNF-α- and IFN-γ-induced HaCaT cells. At the protein level determined by ELISA, WCR significantly reduced the secretion of IL-17A, IL-22, and IL-23. The WCR at low concentrations was proved to possess anti-inflammatory effect without cytotoxicity and it did not interfere with cell cycle of keratinocytes. This is the first study to provide convincing evidence that WCR is a potential candidate for development of effective psoriasis therapies.

Kaempferia parviflora Extract Exhibits Anti-cancer Activity against HeLa Cervical Cancer Cells.

Kaempferia parviflora (KP) has been traditionally used as a folk remedy to treat several diseases including cancer, and several studies have reported cytotoxic activities of extracts of KP against a number of different cancer cell lines. However, many aspects of the molecular mechanism of action of KP remain unclear. In particular, the ability of KP to regulate cancer cell growth and survival signaling is still largely unexplored. The current study aimed to investigate the effects of KP on cell viability, cell migration, cell invasion, cell apoptosis, and on signaling pathways related to growth and survival of cervical cancer cells, HeLa. We discovered that KP reduced HeLa cell viability in a concentration-dependent manner. The potent cytotoxicity of KP against HeLa cells was associated with a dose-dependent induction of apoptotic cell death as determined by flow cytometry and observation of nuclear fragmentation. Moreover, KP-induced cell apoptosis was likely to be mediated through the intrinsic apoptosis pathway since caspase 9 and caspase 7, but not BID, were shown to be activated after KP exposure. Based on the observation that KP induced apoptosis in HeLa cell, we further investigated the effects of KP at non-cytotoxic concentrations on suppressing signal transduction pathways relevant to cell growth and survival. We found that KP suppressed the MAPK and PI3K/AKT signaling pathways in cells activated with EGF, as observed by a significant decrease in phosphorylation of ERK1/2, Elk1, PI3K, and AKT. The data suggest that KP interferes with the growth and survival of HeLa cells. Consistent with the inhibitory effect on EGF-stimulated signaling, KP potently suppressed the migration of HeLa cells. Concomitantly, KP was demonstrated to markedly inhibit HeLa cell invasion. The ability of KP in suppressing the migration and invasion of HeLa cells was associated with the suppression of matrix metalloproteinase-2 production. These data strongly suggest that KP may slow tumor progression and metastasis in patients with cervical cancer. Taken together, the present report provides accumulated evidence revealing the potent anti-cancer activities of Kaempferia parviflora against cervical cancer HeLa cells, and suggests its potential use as an alternative way for cervical cancer prevention and therapy.