Melanoma, an extremely aggressive cancer, causes the most skin cancer-related deaths, due to metastasis to other areas of the body, such as lymph nodes, lungs, liver, brain or bone. It is ...characterized by high levels of matrix metalloproteinase (MMP)-2 and -9 secretions that degrade the extracellular matrix and basement membrane, allowing cancer cells to spread to distal organs. Various cytokines, mitogens, growth factors, inducers and inhibitors control MMP activities. We investigated the roles of these in regulation of MMP-2 and -9 in human melanoma A-2058 cells. Human A-2058 cells were grown in DMEM supplemented with 15% FBS and antibiotics in 24-well tissue culture plates. At near confluence, the cells were washed with PBS and incubated in serum-free media with phorbol 12-myristate 13-acetate (PMA) at 10, 25, 50 and 100 ng/ml; TNF-α and IL-1β at 0.1, 1, 10 and 25 ng/ml; LPS at 10, 25, 50 and 100 µg/ml; epigallocatechin gallate (EGCG) and doxycycline (Dox) at 10, 25, 50 and 100 µM without and with PMA; a nutrient mixture (NM) containing lysine, proline, ascorbic acid and green tea extract without and with PMA at 10, 50, 100, 500 and 1,000 µg/ml; actinomycin D and cyclohexamide at 2 and 4 µM; retinoic acid and dexamethasone at 50 µM. After 24 h the media were removed and analyzed for MMP-2 and MMP-9 by zymography and densitometry. Melanoma A-2058 demonstrated strong expression of MMP-2 and slight expression of MMP-9. PMA at 100 ng/ml showed no effect on MMP-2 secretion but potently upregulated MMP-9 secretion to 400% that of control. TNF-α showed no significant overall effect on expression of MMP-2 but potent dose-dependent increased MMP-9 secretion with 200% of control at 25 ng/ml. IL-1β showed no significant effect on MMP-2 or MMP-9 secretion by A-2058 cells, except at 25 ng/ml where MMP-2 level was reduced by ~40% and MMP-9 secretion ~50%. LPS treatment showed no significant effect on MMP-2 secretion and enhanced MMP-9 secretion up to 25 µg/ml followed by decreased level. EGCG, NM and doxycycline, without and with PMA, downregulated the expression of MMP-2 and MMP-9 in a dose-dependent manner. Actinomycin D, cyclohexamide and retinoic acid had inhibitory effects on MMP-2, while dexamethasone showed slight stimulatory effect on MMP-2 secretion. Our results showed that select cytokines, mitogens and inhibitors modulated A-2058 MMP-2 and MMP-9 expression. They suggest the clinical potential of MMP inhibitors, especially the non-toxic ones, such as the nutrient mixture and its component EGCG in management of melanoma.
The prognosis of hepatocellular carcinoma (HCC) remains dismal despite any treatment. Matrix metalloproteinases (MMPs) have been researched for their role in tumor invasion and metastasis. Various ...cytokines, mitogens, growth factors, inducers, and inhibitors control MMP activities. In this article, we investigated the roles of these in the regulation of MMP-2, -9 secretions in HCC. Human HCC SK-Hep-1 was grown in appropriate media. At near confluence, the cells were washed with phosphate-buffered saline and incubated in serum-free media with PMA; TNF-α, IL-1β; lipopolysaccharide; epigallocatechin gallate (EGCG) and doxycycline (Dox) at various doses with and without PMA; a nutrient mixture (NM) containing lysine, proline, ascorbic acid, and EGCG with and without PMA at; and actinomycin D and cycloheximide at different doses. After 24 hours, the media were removed and analyzed. SK-Hep-1 expressed bands corresponding to MMP-2 and MMP-9. TNF-α showed an insignificant effect on MMP-2 at doses below 25 at which dose MMP-2 was virtually blocked and a moderate dose-dependent effect on MMP-9. Interleukin-1β demonstrated an insignificant effect on MMP-2 at doses below 25 at which dose MMP-2 was completely blocked and enhanced effects on MMP-9. Lipopolysaccharide showed dose-dependent inhibition of MMP-2 and MMP-9. EGCG, Dox, and NM, without and with PMA, downregulated the expression of MMP-2 and MMP-9. Actinomycin D and cycloheximide also had dose-dependent inhibitory effects on MMPs. Our results showed that cytokines, mitogens, and inhibitors modulated SK-Hep-1 MMP-2 and MMP-9 secretion, suggesting the clinical use of especially potent and nontoxic MMP inhibitor as the NM in management of HCC.
Background:
Lyme disease (LD) is a tick-borne infection caused by Borrelia burgdorferi sensu lato. The current therapeutic approach to this disease is limited to antibiotics. However, after their ...administration, about 20% of patients experience delayed onset of this illness manifesting as lingering persistent symptoms.
Methods:
To determine a suitable approach that would help reduce this number, we examined the efficacy of a composition of polyphenolic compounds (baicalein, luteolin, and rosmarinic acid) with fatty acids (monolaurin and cis-2-decenoic acid), and iodine/kelp in a Lyme disease animal model and volunteers.
Results:
The results showed that 4 weeks of dietary intake of this composition reduced the spirochete burden in animal tissues by about 75%. Basic and differential blood parameters did not show significant differences between control animals and the animals fed with this composition. Also, hepatic and renal toxicity markers were not changed and apoptosis was not observed. Relevant inflammatory cytokines such as IL-6, IL-17, TNF-α, and INF-γ, were elevated in infected animals but normalized in infected and treated animals. A small observational study revealed that after administration of this composition to 17 volunteers three times per day for 6 months, 67.4% of the volunteers with late or persistent LD, and not receptive to previous antibiotic application, responded positively, in terms of energy status as well as physical and psychological wellbeing to supplementation with this composition, while 17.7% had slight improvement, and 17.7% were none responsive.
Conclusion:
We concluded that this specific composition revealed feasible benefits in late or persistent LD management, although double-blind controlled clinical trials are warranted.
Ovarian cancer is the deadliest gynecological malignancy in women, and fifth leading cause of death. Despite advances made in chemotherapy and surgery, the average time of clinical remission is ...approximately 2 years and the 5-year survival rate is 45%. Thus, there is an urgent need for the development of a novel therapeutic approach to ovarian cancer treatment. We investigated the effect of a specific nutrient mixture (EPQ) containing ascorbic acid, lysine, proline, green tea extract, and quercetin on human ovarian cancer cell A-2780 in vivo and in vitro. Athymic female nude mice (n = 12) were all inoculated intraperitoneally (IP) with 2 × 10⁶ cells in 0.1 mL of phosphate buffered saline (PBS) and randomly divided into two groups. Upon injection, the Control group (n = 6) was fed a regular diet and the EPQ group (n = 6) a regular diet supplemented with 0.5% EPQ. Four weeks later, the mice were sacrificed and tumors that developed in the ovary were excised, weighed, and processed for histology. Lungs were inspected for metastasis. In vitro, A-2780 cells were cultured in Dulbecco modified Eagle medium supplemented with 10% FBS and antibiotics. At near confluence, cells were treated with EPQ in triplicate at concentrations between 0 and 1000 μg/mL. Cell proliferation was measured via MTT assay, MMP-9 secretion via gelatinase zymography, invasion through Matrigel and morphology via hematoxylin and eosin (H & E) staining. All Control mice developed large ovarian tumors, whereas 5 out of 6 mice in the EPQ group developed no tumors, and one, a small tumor. Control mice also showed lung metastasis in 6 out of 6 mice, while no lung metastasis was evident in EPQ mice. Zymography demonstrated only MMP-9 expression, which EPQ inhibited in a dose-dependent fashion, with virtual total block at 250 μg/mL concentration. EPQ significantly inhibited invasion through Matrigel with total block at 250 μg/mL concentration. MTT showed dose-dependent inhibition of cell proliferation with EPQ, and H & E staining showed no morphological changes below 500 μg/mL EPQ. These results suggest that EPQ has therapeutic potential in the treatment of ovarian cancer by significantly suppressing ovarian tumor incidence and growth and lung metastasis, and by inhibiting MMP-9 secretion and invasion of A-2780 ovarian cancer cells.
Background:
Angiotensin-converting enzyme II or ACE2 is an integral membrane protein present on many types of cells, including vascular endothelial cells and lung alveolar epithelial cells. This ...receptor serves as the entry point for SARS-coronaviruses (SARS-CoVs), including a novel coronavirus 2019-nCoV. Limited availability of these receptors can thwart cellular entry of this virus.
Methods:
We tested the effects of ascorbic acid (vitamin C) on cellular expression of ACE2 at the protein and RNA levels in human small alveolar epithelial cells and microvascular endothelial cells. In addition, we investigated whether combinations of ascorbic acid with other natural compounds can affect ACE2 expression.
Results:
The results show that ascorbic acid itself has moderate but consistent lowering effects on ACE2 expression at the cellular, protein, and RNA levels. Some natural compounds were effective in lowering ACE2 cellular expression, with the highest inhibitory effects observed for baicalin (75%) and theaflavin (50%). Significantly, combinations of these and other test compounds with ascorbic acid further decreased ACE2 expression. The highest impact of ascorbate on ACE2 expression was noted when combined with theaflavin (decrease from 50% to 87%), zinc (decrease from 22% to 62%), and with 10-undecenoic acid (from 18% to 53%). Ascorbic acid showed moderate additional benefits in decreasing ACE2 expression when combined with N-acetylcysteine and baicalin.
Conclusion:
Our study provides valuable experimental confirmation of the efficacy of micronutrients in controlling ACE2 expression—the coronavirus cellular “entry” point. It further validates the importance of nutrient interactions in various aspects of cellular metabolism and in considering potential therapeutic applications of nutrient-based approaches. The study shows that ascorbic acid and its combination with some natural compounds could be included in developing preventive and therapeutic approaches toward the current pandemic.
The ongoing rise in antibiotic resistance, and a waning of the introduction of new antibiotics, has resulted in limited treatment options for bacterial infections, including these caused by ...methicillin-resistant Staphylococcus aureus, leaving the world in a post-antibiotic era. Here, we set out to examine mechanisms by which theaflavin 3,3'-digallate (TF3) might act as an anti-hemolytic compound. In the presented study, we found that TF3 has weak bacteriostatic and bactericidal effects on Staphylococcus aureus, and strong inhibitory effect towards the hemolytic activity of its alpha-hemolysin (Hla) including its production and secretion. A supportive SPR assay reinforced these results and further revealed binding of TF3 to Hla with KD = 4.57x10.sup.-5 M. Interestingly, TF3 was also able to protect human primary keratinocytes from Hla-induced cell death, being at the same time non-toxic for them. Further analysis of TF3 properties revealed that TF3 blocked Hla-prompting immune reaction by inhibiting production and secretion of IL1beta, IL6, and TNFalpha in vitro and in vivo, through affecting NFkappaB activity. Additionally, we observed that TF3 also markedly attenuated S. aureus-induced barrier disruption, by inhibiting Hla-triggered E-cadherin and ZO-1 impairment. Overall, by blocking activity of Hla, TF3 subsequently subdued the inflammation and protected the epithelial barrier, which is considered as beneficial to relieving skin injury.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
The limited ability of current treatments to control metastasis and the proposed antitumor properties of specific nutrients prompted us to examine the effect of a specific formulation (nutrient ...supplement NS) of lysine, proline, arginine, ascorbic acid, and green tea extract in vivo on the development of N-methyl-N-nitrosourea (MNU)-induced mammary tumors in rats.
A single intraperitoneal dose of MNU was injected into each of 20 female Sprague-Dawley rats (aged 50 days) to induce tumors. Two weeks after MNU treatment, a time by which the animals had recovered from MNU-induced toxicity, the rats were divided into two groups. Rats in group 1 (n = 10) were fed Purina chow diet, whereas those in group 2 (n = 10) were fed the same diet supplemented with 0.5% NS. After a further 24 weeks, the rats were killed and tumors were excised and processed.
NS reduced the incidence of MNU-induced mammary tumors and the number of tumors by 68.4%, and the tumor burden by 60.5%. The inhibitory effect of NS was also reflected by decreased tumor weight; the tumor weights per rat and per group were decreased by 41% and 78%, respectively. In addition, 30% of the control rats developed ulcerated tumors, in contrast to 10% in the nutrient supplemented rats.
These findings suggest that the specific formulation of lysine, proline, arginine, ascorbic acid, and green tea extract tested significantly reduces the incidence and growth of MNU-induced mammary tumors, and therefore has strong potential as a useful therapeutic regimen for inhibiting breast cancer development.
Abstract
Current cancer therapies are based on chemotherapy and radiation. These therapeutic approaches can initially reduce the tumor mass or tumor burden by killing tumor cells. It has been ...postulated that the resulting dying tumor cells, or tumor debris, can act as sources of tumor stimulation on surviving cells leading to reappearance of the cancer, and thereby drastically reducing the survival rate. In our study we investigated whether 4T1 debris induced by chemotherapeutic agent docetaxel would affect breast tumor growth in female nude mice. Female athymic nude mice were divided into three groups: Group 1 was inoculated with 4T1 cells; Group 2 with 4T1 cells together with their debris and Group 3 with debris alone. All mice were fed a regular diet. After four weeks, the mice were sacrificed; tumors were excised, weighed and processed for histology and immunohistochemistry for inflammation parameters. Tumor weight significantly increased in Group 2 mice compared to the control Group 1. Group 3 mice developed no tumors. Although the tumor histology for both Groups 1 and 2 were similar, there were significant differences in inflammatory parameters in the two groups. Tumors developed in Group 2 mice had intense staining patterns for TNF-alpha, IL-6, Ki-67, i-Nos and VEGF compared to tumors in the control Group 1. Our results show that dying cells from docetaxel treatment, or tumor debris, had stimulatory effect on growth of breast cancer 4T1 cells in nude mice which was associated with increased inflammation. This study represents a novel method to evaluate the effects of chemotherapy or radiation on surviving tumor cells and to investigate the effect of various inhibitors on the growth recurrence.
Citation Format: M Waheed Roomi, Aleksandra Niedzwiecki, Matthias Rath. Dying tumor cells (tumor debris) stimulate the growth of 4T1 breast cancer tumor in female athymic nude mice abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2172.
Naturally-occurring compounds are acknowledged for their broad antiviral efficacy. Little is however known about their mutual cooperation. Here, we evaluated in vitro efficacy of the defined mixture ...of agents against the RdRp complex of the original SARS-CoV-2 and Omicron variant. This composition of vitamin C, N-acetylcysteine, resveratrol, theaflavin, curcumin, quercetin, naringenin, baicalin, and broccoli extract showed to inhibit activity of RdRp/nsp7/nsp8 both these variants. In vitro exposure of recombinant RdRp complex to individual compounds of this composition pointed to quercetin as the driving inhibitory compound. The outcome of this study supports the motion of antiviral efficacy of natural compounds against SARS-CoV-2 and Omicron and implies that their reciprocal or mutual interaction may augment antiviral action through simultaneous effect on different mechanisms. Consequently, this makes it more difficult for an infectious agent to evade all these mechanisms at the same time. Considering the urgency in finding effective prevention, but also side-effects free treatment of COVID-19 our results call for clinical affirmation of the benefits of this micronutrient combination in both preventive and therapeutic aspects. Whether observed effects can be achieved, by concentrations of the active agents used in these in vitro experiments, in in vivo or clinical setting warrants further study.
Borrelia sp. is a causative pathogen of Lyme disease which has become a worldwide health concern. Non-toxic approaches especially directed toward latent persistent forms of this pathogen are desired. ...Lipids in the form of volatile and non-volatile oils, and fatty acids with proven anti-borreliae efficacy could become an additional support or an alternative for consideration in treatment approaches.
In this study we investigated 47 lipids (30 volatile and non-volatile oils, and 17 fatty acids) of plant and animal origin against typical motile, knob/round-shaped persisters, and biofilm-like aggregates of Borrelia burgdorferi s.s. and Borrelia garinii, which are identified as pathogenic factors of Lyme disease in the USA and Europe, using direct microscopic counting and spectrofluorometric measurements.
Out of all examined lipids, 5 oils (Bay leaf oil, Birch oil, Cassia oil, Chamomile oil German, and Thyme oil) at or below 0.25%, and 3 fatty acids (13Z,16Z Docosadienoic acid, erucic acid, and petroselinic acid) at or below 0.75 mg/ml, showed bactericidal activity against typical motile spirochetes and knob/round-shaped persisters. Only Bay leaf oil and Cassia oil, including their major constituents, eugenol and cinnamaldehyde, showed to target biofilm-like aggregates of both tested Borrelia spp. at the same concentration, although with 20-30% eradication mark.
Based on obtained results, volatile oils were more potent than non-volatile oils, and unsaturated fatty acids were more effective than saturated fatty acids. Among all tested oils, Bay leaf oil and Cassia oil, with their major components eugenol and cinnamaldehyde, seem to have the highest anti-borreliae efficacy.
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![Inhibition of [alpha]-hemol...](http://api.summon.serialssolutions.com/2.0.0/image/issn/XR4PS5YY4K/1932-6203/small "Inhibition of [alpha]-hemolysin activity of Staphylococcus aureus by theaflavin 3,3'-digallate")
