Remodeling of Cancer-Specific Metabolism under Hypoxia with Lactate Calcium Salt in Human Colorectal Cancer Cells.

Hypoxic cancer cells meet their growing energy requirements by upregulating glycolysis, resulting in increased glucose consumption and lactate production. Herein, we used a unique approach to change in anaerobic glycolysis of cancer cells by lactate calcium salt (CaLac). Human colorectal cancer (CRC) cells were used for the study. Intracellular calcium and lactate influx was confirmed following 2.5 mM CaLac treatment. The enzymatic activation of lactate dehydrogenase B (LDHB) and pyruvate dehydrogenase (PDH) through substrate reaction of CaLac was investigated. Changes in the intermediates of the tricarboxylic acid (TCA) cycle were confirmed. The cell viability assay, tube formation, and wound-healing assay were performed as well as the confirmation of the expression of hypoxia-inducible factor (HIF)-1α and vascular endothelial growth factor (VEGF). In vivo antitumor effects were evaluated using heterotopic and metastatic xenograft animal models with 20 mg/kg CaLac administration. Intracellular calcium and lactate levels were increased following CaLac treatment in CRC cells under hypoxia. Then, enzymatic activation of LDHB and PDH were increased. Upon PDH knockdown, α-ketoglutarate levels were similar between CaLac-treated and untreated cells, indicating that TCA cycle restoration was dependent on CaLac-mediated LDHB and PDH reactivation. CaLac-mediated remodeling of cancer-specific anaerobic glycolysis induced destabilization of HIF-1α and a decrease in VEGF expression, leading to the inhibition of the migration of CRC cells. The significant inhibition of CRC growth and liver metastasis by CaLac administration was confirmed. Our study highlights the potential utility of CaLac supplementation in CRC patients who display reduced therapeutic responses to conventional modes owing to the hypoxic tumor microenvironment.

Development of Glycerol-Rose Bengal-Polidocanol (GRP) foam for enhanced sclerosis of a cyst for cystic diseases.

Polycystic kidney disease (PKD) is a common genetic disorder that results in a proliferating and enlarging cyst and ultimately leads to loss of kidney function. Because an enlarged cyst is a primary factor for limited kidney function, the large cyst is surgically removed by laparoscopic deroofing or sclerosant. This a relatively nascent treatment method entails complications and sometimes fail due to the cyst fluid refilling and infection. This study proposes using a more stable and effective polidocanol foam with glycerol and Rose Bengal (GRP form) to prevent cyst regeneration and irritation, which is caused by the required body movement during the treatment. Specifically, the foam retention time and viscosity were increased by adding glycerol up to 10% (w/v). The GRP form inhibited cellular proliferation and disrupted cellular junctions, e-cadherin, and cyst formation, demonstrated by the LDH, Live and Dead, and re-plating culture assays. The GRP foam was shown to be a safe and effective treatment as a commercial grade polidocanol foam form by an in vivo study in which subcutaneously injected mice injected with commercial 3% polidocanol, and the GRP foam showed no difference in inflammation. Thus, this study provides an advanced polidocanol form by adding glycerol and Rose-Bengal to help existing sclerotherapy.

FlexPro MD®, a Combination of Krill Oil, Astaxanthin and Hyaluronic Acid, Reduces Pain Behavior and Inhibits Inflammatory Response in Monosodium Iodoacetate-Induced Osteoarthritis in Rats.

Osteoarthritis (OA) is a degenerative joint disease and a leading cause of adult disability. Since there is no cure for OA and no effective treatment to slow its progression, current pharmacologic treatments, such as analgesics and non-steroidal anti-inflammatory drugs (NSAIDs), only alleviate symptoms, such as pain and inflammation, but do not inhibit the disease process. Moreover, chronic intake of these drugs may result in severe adverse effects. For these reasons, patients have turned to the use of various complementary and alternative approaches, including diverse dietary supplements and nutraceuticals, in an effort to improve symptoms and manage or slow disease progression. The present study was conducted to evaluate the anti-osteoarthritic effects of FlexPro MD® (a mixture of krill oil, astaxanthin, and hyaluronic acid; FP-MD) in a rat model of OA induced by monosodium iodoacetate (MIA). FP-MD significantly ameliorated joint pain and decreased the severity of articular cartilage destruction in rats that received oral supplementation for 7 days prior to MIA administration and for 21 days thereafter. Furthermore, FP-MD treatment significantly reduced serum levels of the articular cartilage degeneration biomarkers cartilage oligomeric matrix protein (COMP) and crosslinked C-telopeptide of type II collagen (CTX-II), and the pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α), interleukin-1ß (IL-1ß), and interleukin-6 (IL-6), as well as mRNA expression levels of inflammatory mediators, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), and matrix-degrading enzymes, matrix metalloproteinase (MMP)-2 and MMP-9, in the knee joint tissue. Our findings suggest that FP-MD is a promising dietary supplement for reducing pain, minimizing cartilage damage, and improving functional status in OA, without the disadvantages of previous dietary supplements and medicinal agents, including multiple adverse effects.

N,N'-Diacetyl-p-phenylenediamine restores microglial phagocytosis and improves cognitive defects in Alzheimer's disease transgenic mice.

As a central feature of neuroinflammation, microglial dysfunction has been increasingly considered a causative factor of neurodegeneration implicating an intertwined pathology with amyloidogenic proteins. Herein, we report the smallest synthetic molecule (N,N'-diacetyl-p-phenylenediamine [DAPPD]), simply composed of a benzene ring with 2 acetamide groups at the para position, known to date as a chemical reagent that is able to promote the phagocytic aptitude of microglia and subsequently ameliorate cognitive defects. Based on our mechanistic investigations in vitro and in vivo, 1) the capability of DAPPD to restore microglial phagocytosis is responsible for diminishing the accumulation of amyloid-ß (Aß) species and significantly improving cognitive function in the brains of 2 types of Alzheimer's disease (AD) transgenic mice, and 2) the rectification of microglial function by DAPPD is a result of its ability to suppress the expression of NLRP3 inflammasome-associated proteins through its impact on the NF-κB pathway. Overall, our in vitro and in vivo investigations on efficacies and molecular-level mechanisms demonstrate the ability of DAPPD to regulate microglial function, suppress neuroinflammation, foster cerebral Aß clearance, and attenuate cognitive deficits in AD transgenic mouse models. Discovery of such antineuroinflammatory compounds signifies the potential in discovering effective therapeutic molecules against AD-associated neurodegeneration.

Characterization of the Subventricular-Thalamo-Cortical Circuit in the NP-C Mouse Brain, and New Insights Regarding Treatment.

Gliosis in Niemann-Pick type C (NP-C) disease is characterized by marked changes in microglia and astrocytes. However, the gliosis onset and progression in NP-C has not been systematically studied, nor has the mechanism underlying this finding. Here, we found early gliosis in the subventricular zone (SVZ) of NP-C mice. Neural progenitor damage by Npc1 mutation suppressed vascular endothelial growth factor (VEGF) expression and further induced microglia activation followed by astrogliosis. Interestingly, excessive astrogliosis in the SVZ induced neural progenitor retention and/or migration into thalamus via astrocyte-derived VEGF, resulting in acceleration of thalamic and cortical gliosis through thalamo-cortical pathways. Transplantation of VEGF-overexpressing neural stem cells into the SVZ improved whole-brain pathology of NP-C mice. Overall, our data provide a new pathological perspective on NP-C neural pathology, revealing abnormalities in the subventricular-thalamo-cortical circuit of NP-C mouse brain and highlighting the importance of the SVZ microenvironment as a therapeutic target for NP-C disease.

CCL2 induces neural stem cell proliferation and neuronal differentiation in Niemann-Pick type C mice.

Niemann-Pick type C disease (NP-C) is a rare and ultimately fatal lysosomal storage disorder with variable neurologic symptoms. Loss of neuronal function and neuronal cell death occur in the NP-C brain, similar to the findings for other neurodegenerative diseases. Targeting of neuronal cells in the brain therefore represents a potential clinical intervention strategy to reduce the rate of disease progression and improve the quality of life. We previously reported that bone marrow stem cells show a neurogenic effect through CCL2 (also known as monocyte chemoattractant protein-1, MCP-1) secretion in the brains of NP-C mice. However, the direct effect of CCL2 on neurogenesis has not been ascertained. Here, to define neurogenic effects of CCL2 in NP-C, we applied human recombinant CCL2 to neural stem cells (NSCs) derived from NP-C mice. CCL2-treated NSCs showed significantly increased capacity for self-renewal, proliferation and neuronal differentiation. Similar results were observed in the subventricular zone of NP-C mice after CCL2 treatment. Furthermore, infusion of CCL2 into the NP-C mouse brain resulted in reduction of neuroinflammation. Taken together, our results demonstrate that CCL2 is a potential new therapeutic agent for NP-C.

Recombinant soluble neprilysin reduces amyloid-beta accumulation and improves memory impairment in Alzheimer's disease mice.

Accumulation of amyloid-ß (Aß) is thought to be a central pathology in the brain of patients with Alzheimer's disease (AD). Neprilysin (NEP), a plasma membrane glycoprotein of the neutral zinc metalloendopeptidase family, is known as a major Aß-degrading enzyme in the brain. The level of NEP is reduced in the brains of patients with AD; therefore, NEP is under intense investigation as a potential therapeutic source for degradation of deposited Aß in AD. Previous studies have utilized viral vectors expressing NEP for reduction of Aß deposition in the brain. However, viral vectors have disadvantages regarding difficulty in control of insert size, expression desired (short- or long-term), and target cell type. Here, in order to overcome these disadvantages, we produced recombinant soluble NEP from insect cells using an NEP expression vector, which was administered by intracerebral injection into AD mice, resulting in significantly reduced accumulation of Aß. In addition, AD mice treated with NEP showed improved behavioral performance on the water maze test. These data support a role of recombinant soluble NEP in improving memory impairment by regulation of Aß deposition and suggest the possibility that approaches using protein therapy might have potential for development of alternative therapies for treatment of AD.

Effects of roasting conditions on the physicochemical properties and volatile distribution in perilla oils (Perilla frutescens var. japonica).

UNLABELLED: Perilla seeds have more than 60% of α-linolenic acid, one of omega-3 essential fatty acids. Headspace volatiles and physicochemical properties including color, fluorescence intensity, and the oxidation products in perilla oil (PO) from perilla seeds roasted at different conditions were analyzed. Roasting temperature was 150, 170, 190, and 210°C, and roasting time was 15 and 30 min at each roasting temperature. PO from higher roasting temperature and longer roasting time had lower L* values, higher a*, b*, and chroma values, more brown pigments and fluorescence intensity, and more conjugated dienoic acids. Pyrazines were major volatiles in PO, and furans, sulfur-containing compounds, and hydrocarbons were also detected by a solid phase microextraction gas chromatography/mass spectrometry. In PO, 2,5-Dimethylpyrazine and 2-furancarboxaldehyde were 2 major volatiles. The principal component analysis of volatiles showed the 1st principal component (PC1) and the 2nd principal component (PC2) express 56.64% and 22.72% of the volatile variability in PO, respectively, which can differentiate PO prepared from roasting conditions clearly. Some physicochemical properties especially brown pigment and volatiles were positively correlated with each other in PO. PRACTICAL APPLICATION: Perilla oil (PO) from perilla seeds possesses more than 60% of α-linolenic acid, one of omega-3 fatty acids. Roasting process has been used to extract oil from perilla seeds. Understanding physicochemical properties of PO from diverse roasting conditions are important steps to produce PO in food industry. Roasting process induces darkening of color, increase of fluorescence intensity, and brown pigments in PO. Pyrazines and furans are major headspace volatiles in PO roasted above 170°C. The results of this study can help to produce PO in industrial scales with desired headspace volatiles, colors, and oxidative state.

Application of solid phase-microextraction (SPME) and electronic nose techniques to differentiate volatiles of sesame oils prepared with diverse roasting conditions.

Headspace volatiles of sesame oil (SO) from sesame seeds roasted at 9 different conditions were analyzed by a combination of solid phase microextraction (SPME)-gas chromatography/mass spectrometry (GC/MS), electronic nose/metal oxide sensors (MOS), and electronic nose/MS. As roasting temperature increased from 213 to 247 °C, total headspace volatiles and pyrazines increased significantly (P < 0.05). Pyrazines were major volatiles in SO and furans, thiazoles, aldehydes, and alcohols were also detected. Roasting temperature was more discrimination factor than roasting time for the volatiles in SO through the principal component analysis (PCA) of SPME-GC/MS, electronic nose/MOS, and electronic nose/MS. Electronic nose/MS showed that ion fragment 52, 76, 53, and 51 amu played important roles in discriminating volatiles in SO from roasted sesame seeds, which are the major ion fragments from pyrazines, furans, and furfurals. SO roasted at 213, 230, and 247 °C were clearly differentiated from each other on the base of volatile distribution by SPME-GC/MS, electronic nose/MOS, and electronic nose/MS analyses. Practical Application: The results of this study are ready to apply for the discriminating samples using a combinational analysis of volatiles. Not only vegetable oils prepared from roasting process but also any food sample possessing volatiles could be targets for the SPME-GC/MS and electronic nose assays. Contents and types of pyrazines in sesame seed oil could be used as markers to track down the degree of roasting and oxidation during oil preparation.

Effects of riboflavin photosensitization on the degradation of bisphenol A (BPA) in model and real-food systems.

Effects of riboflavin photosensitizations on the stability of bisphenol A (BPA), a well-known endocrine disrupting chemical, were studied in model and real-food systems by high-performance liquid chromatography (HPLC). Concentration of BPA was significantly decreased under light exposure (P < 0.05) as the concentration of riboflavin increased while those without riboflavin under light or those with riboflavin in the dark did not change significantly (P > 0.05). Addition of 50, 100, and 200 microM sodium azide significantly increased the stability of BPA in riboflavin photosensitization with concentration dependent manner (P < 0.05), implying that a singlet oxygen or type II pathway played a role in the photodegradation of BPA. Stability of BPA in riboflavin was significantly increased in the presence of high concentration of tert-butanol, a hydroxyl radical quencher, under light storage for 80 min, indicating hydroxyl radicals were involved and contributed to the degradation of BPA, at least in part. Availability of riboflavin photosensitization on the photodegradation of BPA was tested in 2 canned tea beverages with different phenolic contents. BPA was more stable in the beverage sample with higher total phenolic contents and free radical scavenging ability. The photodegradation of BPA in riboflavin photosensitization can be an efficient way to decrease the concentration of BPA from environmental or food systems.

Growth and rutin production in hairy root cultures of buckwheat (Fagopyrum esculentum M.).

Buckwheat (Fagopyrum esculentum Moench.) is a potentially important source of rutin, a natural flavonoid with antihyperglycemic, antihypertensive, and antioxidative properties. To examine in vitro production of rutin, we established a hairy root culture of buckwheat by infecting leaf explants with Agrobacterium rhizogenes R1000, and tested the growth conditions and rutin production rates of these cultures. Ten hairy root clones were established; their growth and rutin production rates ranged from 233 to 312 (mg dry wt per 30 mL flask, and 0.8 to 1.2 (mg/g dry wt), respectively. Clone H8, which had high growth and rutin production rates (312 mg dry wt per 30 mL flask and 1.2 mg/g dry wt, respectively), was selected for further experiments. H8 showed maximal growth and rutin content at 30 days in culture in MS medium. Of four tested culture media, half-strength MS medium was found to induce the highest levels of growth (378 mg dry wt per 30 mL flask) and rutin production (1.4 mg/g dry wt) by clone H8. In contrast, supplementation with auxins (0.1-1 mg/l IAA, IBA and NAA) increased the growth rate, but had no significant effect on rutin production by H8. Collectively, these findings indicate that hairy root cultures of buckwheat culture could be a valuable alternative approach for rutin production.