Gustatory signaling begins with taste receptor cells that express taste receptors. Recent molecular biological studies have identified taste receptors and transduction components for basic tastes (sweet, salty, sour, bitter, and umami). Activation of these receptor systems leads to depolarization and an increase in [Ca2+]i in taste receptor cells. Then transmitters are released from taste cells and activate gustatory nerve fibers. The connection between taste cells and gustatory nerve fibers would be specific because there may be only limited divergence of taste information at the peripheral transmission. Recent studies have demonstrated that sweet taste information can be modulated by hormones or other endogenous factors that could act on their receptors in a specific group of taste cells. These peripheral modulations of taste information may influence preference behavior and food intake. This paper summarizes data on molecular mechanisms for detection and transduction of taste signals in taste bud cells, information transmission from taste cells to gustatory nerve fibers, and modulation of taste signals at peripheral taste organs, in particular for sweet taste, which may play important roles in regulating energy homeostasis.
Afferent signals from the olfactory system, gustatory system and gastrointestinal (GI) tract control visceral functions such as oral and gut secretions and several digestive, endocrine, thermogenic, cardiovascular and renal responses via autonomic reflexes. It is well known that odors and tastes, such as umami, can stimulate oral and GI secretions to improve food intake and digestion in a process termed the cephalic phase response. The perception of GI nutrients, such as carbohydrates and amino acids, can control food digestion, absorption and utilization via the vago-vagal reflex during a meal. Recent advances in understanding the molecular physiology of taste indicate that taste receptors able to sense such nutrients are widely distributed in the GI tract, including the oral cavity. These receptors act as nutrient sensors to trigger food digestion, the release of GI peptides and the formation of food preferences. In this paper, we review recent evidence on the regulation of GI functions by the autonomic nervous system via peripheral odor and nutrient sensors.
Aspartate, an excitatory amino acid, is known to be stored in synaptic vesicles and exocytosed from some neurons to perform aspartergic neurotransmission. Through in vitro reconstitution, we found that sialin, a lysosomal sialic acid exporter, is responsible for the vesicular storage of aspartate in hippocampal neurons and pinealocytes. Mutations found in Salla disease cause decreased aspartate transport activity without affecting sialic acid transport. Thus, sialin is a multifunctional transporter. It is possible that people with Salla disease lose the ability of aspartergic neurotransmission, and this could explain why Salla disease involves severe neurological defects.
Swallowing involves several motor processes such as bolus formation and intraoral transport of a food bolus (oral stage) and a series of visceral events that occur in a relatively fixed timed sequence but are to some degree modifiable (pharyngeal stage or swallow reflex). Reflecting the progressive aging of society, patients with swallowing disorders (i.e., dysphagia) are increasing. Therefore, there is expanding social demand for the development of better rehabilitation treatment of dysphagic patients. To date, many dysphagia diets have been developed and are available commercially to help bring back the pleasure of mealtimes to dysphagia patients. Texture modification of food to make the food bolus easier to swallow with less risk of aspiration is one of the important elements in dysphagia diets from the viewpoint of safety assurance. However, for the further development of dysphagia diets, new attempts based on new concepts are needed. One of the possible approaches is to develop dysphagia diets that facilitate swallow initiation. For this approach, an understanding of the mechanisms of swallow initiation and identification of factors that facilitate or suppress swallow initiation are important. In this review, we first summarize the neural mechanisms of swallowing and effects of taste and other inputs on swallow initiation based on data mainly obtained from experimental animals. Then we introduce a recently established technique for eliciting swallowing using electrical stimulation in humans and our ongoing studies using this technique.
Enjoying taste should be one of the greatest pleasures in human life. However, aging is sometimes associated with decreased taste sensitivity, also known as hypogeusia. The loss of taste not only affects quality of life, but can also cause weight loss and health problems in the elderly. Our recent study has shown that 37% of test subjects over 65 years of age exhibited hypogeusia. Further, whole saliva secretion, including minor salivary secretion, was significantly decreased in elderly patients with gustatory impairment, but was normal in all elderly subjects with normal taste thresholds. These data indicate that hyposalivation is closely related to hypogeusia. Moreover, clinical studies have shown that treatment of hyposalivation diminishes hypogeusia, indicating that salivation is essential to maintain normal taste function. However, many medications for relief of dry mouth, such as parasympathomimetic (cholinomimetic) drugs, have serious adverse effects. Palpitation, sweating, nausea, diarrhea and dizziness have all been observed in elderly patients taking parasympathomimetic drugs. To circumvent this problem, glutamate, which produces umami taste, was demonstrated to increase salivary secretion and thereby improve hypogeusia by enhancing the gustatory-salivary reflex. Our data suggests that umami is an effective tool for the relief of hypogeusia without the side effects of parasympathomimetic drugs.
Some ingested nutrients have postingestive effects that modulate food intake and improve mood subconsciously. Here, we provide an overview of the positive postingestive effects of such nutrients, primarily L-glutamate, sugar, and lipids, with respect to behavior and brain function. We also discuss the mechanisms of brain activation resulting from signaling through the gut-brain axis. Recent studies have shown that rats prefer solutions paired with intragastric nutrients that have positive postingestive effects. Using functional magnetic resonance imaging (fMRI), we previously evaluated neural activation in response to ingested glucose, L-glutamate, and corn oil emulsion in rats and found that distinct forebrain regions were activated by these nutrients. Most of the areas activated by intragastric administration of L-glutamate were eliminated by abdominal vagotomy. On the other hand, the areas activated by intragastric administration of glucose were not affected by vagotomy. A behavioral study showed similar results for L-glutamate and glucose. These results indicate that brain activation in response to ingested nutrients occurs through distinct internal signals from the gut to the brain. Distinct regional and temporal activation in the brain determines the variety of postingestive behaviors and physiological responses.
The activity and processing of mammalian S-adenosylmethionine decarboxylase (AdoMetDC) is stimulated by putrescine. To obtain new insights into the mechanism through which putrescine stimulates AdoMetDC, we investigated conformational changes in rat prostate AdoMetDC in the presence or absence of putrescine. We examined the reactivity of purified rat prostate AdoMetDC to the SH-reagent iodoacetic acid (IAA) and its susceptibility to proteolysis in the presence or absence of putrescine using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). The activity of AdoMetDC treated with IAA in the absence of putrescine was reduced, but about 80% of its activity remained after treatment with IAA in the presence of putrescine. In the presence of putrescine, IAA incorporation was 1.9 mol IAA/mol of AdoMetDC α-subunit, while there was no incorporation of IAA in the β-subunit of AdoMetDC. In the absence of putrescine, 5.0 mol of IAA/mol of α-subunit and 0.9 mol of IAA/mol of β-subunit were incorporated. Only Cys292 and Cys310 were carboxymethylated by IAA in the presence of putrescine. In contrast, in the absence of putrescine all cysteines were carboxymethylated by IAA. In addition, putrescine slowed the rate of AdoMetDC degradation by trypsin. These results demonstrate that the conformation of AdoMetDC purified from rat prostate is stabilized by putrescine.
In this study, we examined the substrate specificity, inhibitor sensitivity and kinetic mechanism of a rat aldose reductase-like protein, which is named AKR1B14 in the aldo-keto reductase (AKR) superfamily. AKR1B14 catalyzed the nicotinamide adenine dinucleotide phosphate reduced form (NADPH)-dependent reduction of carbonyl compounds (derived from lipid peroxidation and glycation), xenobiotic aromatic aldehydes and some aromatic ketones. 4-Oxo-2-nonenal, the best substrate showing a Km value of 0.16 μM, was reduced into less reactive 4-oxo-2-nonenol, and its cytotoxicity was attenuated by the overexpression of the enzyme in cultured cells. The enzyme also showed low Km values (0.9—10 μM) for medium-chain aliphatic aldehydes (such as 4-hydroxynonenal, 1-hexenal and farnesal) and 3-deoxyglucosone, although the Km values for short-chain substrates (such as isocaproaldehyde, acrolein and methylglyoxal) were high (16—600 μM). In the reverse reaction, aliphatic and aromatic alcohols were oxidized by AKR1B14 at low rates. AKR1B14 was inhibited by aldose reductase inhibitors such as tolrestat and epalrestat, and their inhibition patterns were noncompetitive versus the aldehyde substrate and competitive with respect to the alcohol substrate. Kinetic analyses of the oxidoreduction and dead-end inhibition suggest that the reaction follows an ordered sequential mechanism.
Endothelial dysfunction/loss is a key event in the development of vascular diseases, including native atherosclerosis (AS). Recent studies have shown that endothelial progenitor cells (EPCs) have the ability to repair endothelial cells that have been lost or damaged following AS. As a result, the therapy of transplanting EPCs is a promising option for the treatment of AS. However, the therapeutic effect on AS with only EPCs transplantation has not been satisfactory. The upregulation of those genes, which prevent the progress of AS in EPCs, is a novel therapeutic strategy for AS. Because it can reduce macrophage foam cell formation and protect endothelial cells from the oxidation of low-density lipoprotein (ox-LDL), paraoxonase-1 (PON1) gene is a candidate for gene therapy in AS. In this study, a recombinant adeno-associated virus (rAAV) vector carrying the human paraoxonase-1 (hPON1) gene (rAAV-PON1) was constructed, and endothelial progenitor cells (EPCs) transfected with rAAV-PON1 were transplanted into the atherosclerosis model of Sprague-Dawley rats (SD rats). The results of doppler ultrasound and histological analysis showed that the group transplanted with the hPON1 gene-transfected EPCs was superior to the group transplanted only with the EPCs and was also better than the group with hPON1 gene injection alone. The results indicated that rAAV-mediated hPON1 gene-transfected EPCs is a potentially valuable new tool in the treatment of atherosclerosis.
A growing body of evidence suggests that nobiletin (5,6,7,8,3′,4′-hexamethoxy flavone) from the peel of citrus fruits, enhances the damaged cognitive function in disease animal models. However, the neuroprotective mechanism has not been clearly elucidated. Since nobiletin has shown anti-inflammatory effects in several tissues, we investigated whether nobiletin suppresses excessive microglial activation implicated in neurotoxicity in lipopolysaccharide (LPS)-stimulated BV-2 microglia cell culture models. Release of nitric oxide (NO), the major inflammatory mediator in microglia, was markedly suppressed in a dose-dependent manner following nobiletin treatment (1—50 μM) in LPS-stimulated BV-2 microglia cells. The inhibitory effect of nobiletin was similar to that of minocycline, a well-known microglial inactivator. Nobiletin significantly inhibited the release of the pro-inflammatory cytokine tumor necrosis factor (TNF-α) and interleukin-1β (IL-1β). LPS-induced phosphorylations of extracellular signal-regulated kinase (ERK), c-Jun NH2-terminal kinase (JNK), and p38 mitogen-activated protein kinases (MAPKs) were also significantly inhibited by nobiletin treatment. In addition, nobiletin markedly inhibited the LPS-induced pro-inflammatory transcription factor nuclear factor κB (NF-κB) signaling pathway by suppressing nuclear NF-κB translocation from the cytoplasm and subsequent expression of NF-κB in the nucleus. Taken together, these results may contribute to further exploration of the therapeutic potential and molecular mechanism of nobiletin in relation to neuroinflammation and neurodegenerative diseases.
Small noncoding microRNAs (miRNAs) have been shown to play an important role in tumor proliferation and metastasis. However, their function and mechanism in the proliferation and metastasis of gastric cancer has not yet been elucidated. Here, we investigated the relationship between miRNA-199a and gastric cancer proliferation and metastasis. Using real-time reverse-transcriptase (RT)-polymerase chain reaction, we found that miR-199a is highly expressed in gastric cancer compared to normal gastric tissues and in metastatic, compared to non-metastatic gastric cancer tissues. MiR-199a positively regulated gastric cancer cell proliferation, migration and invasion. Further studies showed that mitogen-activated protein kinase kinase kinase 11 was significantly down-regulated by miR-199a at the post-transcriptional level and, the level of miR-199a expression in gastric cancer significantly correlated with clinical progression. These findings suggested miR-199a promoted proliferation and metastasis of gastric cancer cells through a regulatory pathway in gastric cancer that has yet to be described. miR-199a may be useful as a new potential therapeutic target for gastric cancer.
Tanshinone IIA (Tan IIA; 14,16-epoxy-20-nor-5(10),6,8,13,15-abietapentaene-11,12-dione), a phytochemical derived from the roots of Salvia miltiorrhiza BUNGE, has been reported to posses anti-angiogenic, anti-oxidant, anti-inflammatory and apoptotic activities. However, the cancer growth inhibitory/cytocidal effects and molecular mechanisms in prostate cancer cells have not been well studied. In the present study, we demonstrate that Tan IIA significantly decreased the viable cell number of LNCaP (phosphate and tensin homolog (PTEN) mutant, high AKT, wild type p53) prostate cancer cells more sensitively than against the PC-3 (PTEN null, high AKT, p53 null) prostate cancer cells. Tan IIA significantly increased TdT-mediated dUTP nick-end labeling (TUNEL) positive index and sub-G1 DNA contents of treated cells, consistent with apoptosis. Tan IIA treatment led to cleavage activation of pro-caspases-9 and 3, but not pro-caspase-8, and cleavage of poly (ADP ribose) polymerase (PARP), a caspase-3 substrate. Additionally, Tan IIA treatment induced cytochrome c release from the mitochondria into the cytosol and reduced mitochondrial membrane potential and suppressed the expression of mitochondria protective Bcl-2 family protein Mcl-1L. Tan IIA reduced the expression of phosphoinositide 3-kinase (PI3K) p85 subunit, and the phosphorylation of AKT and mammalian target of rapamycin (mTOR) in a concentration-dependent manner. Moreover, the combination of Tan IIA and LY294002, a specific PI3K inhibitor, enhanced PARP cleavage of LNCaP and PC-3, but not in MDA-MB-231 breast cancer cells which do not contain detectable active AKT. The findings suggest that Tan IIA-induced apoptosis involves mitochondria intrinsic caspase activation cascade and an inhibition of the PI3K/AKT survival pathway.
Hypoxia is the hallmark of solid tumors and contributes to tumor angiogenesis mainly through activation of the transcription factor hypoxia-inducible factor-1 (HIF-1). In addition to upregulating vascular endothelial growth factor (VEGF) in angiogenesis, HIF-1 plays critical roles in the metabolism, proliferation, metastasis, and differentiation of cancer cells. We and others have previously shown that 1,2,3,4,6-penta-O-galloyl-beta-D-glucose (PGG) from Oriental herbal medicine possesses anti-angiogenic, anti-tumorigenic, and anti-diabetic activities. In the present study, we report that PGG inhibits hypoxia-induced protein accumulation, transcriptional activation, and mRNA expression of HIF-1α in LNCaP prostate cancer cells. PGG reduced cellular and secreted VEGF levels as well as mRNA expression in LNCaP cells. PGG suppressed capillary tube formation in human umbilical vein endothelial cells (HUVECs) maintained in conditioned medium of hypoxia-induced LNCaP cells, indicating that PGG has anti-angiogenic activity under hypoxic condition. Furthermore, PGG reduced expression of phosphoinositide 3-kinase (PI3K) as well as phosphorylation of AKT and mammalian target of rapamycin (mTOR), but not extracellular signal-regulated kinase (ERK) in LNCaP cells under hypoxic condition. Consistently, LY294002, a specific PI3K inhibitor, enhanced the inactivation of HIF-1α and AKT by PGG in LNCaP cells. Taken together, our results demonstrate that PGG inhibits hypoxia-mediated accumulation of HIF-1α as well as its downstream signaling to VEGF and PI3K/AKT/mTOR pathway in LNCaP prostate cancer cells.
Oxidative stress in brain is emerging as a potential causal factor in aging and age-related neurodegenerative disorders. A large body of evidence shows that induction of endogenous antioxidative proteins seems to be a reasonable strategy for delaying the progression of cell injury. In this study, geniposide upregulates the expression of heme oxygenase-1 (HO-1) to attenuate the cell apoptosis induced by 3-morpholinosydnonimine hydrochloride (SIN-1) in primary cultured hippocampal neurons. Furthermore, geniposide induces the nuclear translocation of nuclear factor-E2-related factor 2 (Nrf2) and activation of phosphatidylinositol 3′-kinase (PI3K) in the presence of oxidative stress, and both LY294002 (a specific inhibitor of PI3K) and Zinc protoporphyrin (ZnPP, an inhibitor of HO-1) decrease the cytoprotective action of geniposide in hippocampal neurons. Taken together, the novel cytoprotective mechanism of geniposide to antagonize oxidative stress may be involved in PI3K- and Nrf2-mediated upregulation of the antioxidative enzyme HO-1.
Biglycan (Bgn) is a member of the small leucine-rich proteoglycan (SLRP) family found in bone extracellular matrix (ECM), and hence involved in regulating bone formation and matrix mineralization. It has been reported that Bgn facilitates osteoblast differentiation, and extracellular signal-regulated kinase (Erk) and Smad are two important pathways in regulating osteoblast differentiation. However, the underlying mechanism for Bgn facilitating osteoblast differentiation has not been fully elucidated. The present study demonstrated that the matrix protein Bgn activates Erk signaling pathway and therefore increases Runx2 transcriptional activity, in which glycosaminoglycans (GAGs) chains play an essential role. Additionally, Bgn also activated Smad pathway, another signaling pathway related with osteoblast differentiation. The activation of these two signaling pathways induced by Bgn facilitated the mineralization deposition in vitro. These results demonstrated the mechanism of Bgn promoting osteoblast differentiation and matrix mineralization.
Tenascin-X (TNX), which has a molecular mass of roughly 450 kDa, is the largest member of the tenascin family. Complete deficiency of TNX in humans leads to a recessive form of Ehlers–Danlos syndrome (EDS). TNX is expressed abundantly in a variety of tissues, especially in cardiac muscle and in perivascular stroma. Human TNX is also present in serum with an apparent molecular size of 140 kDa. In the present study, we investigated the expression levels of TNX protein in thoracic and abdominal aortic aneurysm tissues. The level of TNX was significantly increased in both aortic aneurysm tissues compared with that in adjacent normal tissues. Next, to compare TNX levels in serum from both patients with thoracic aortic aneurysm and patients with abdominal aortic aneurysm with levels in serum from healthy individuals, we developed a sandwich enzyme-linked immunosorbent assay (ELISA) using TNX-specific antibodies. Measurement of TNX serum concentrations in both aortic aneurysm patients and controls showed that the levels were almost the same. These results indicate that TNX expression is significantly elevated in both thoracic and abdominal aortic aneurysm tissues but that the increase in TNX levels in both tissues does not result in an increase in TNX serum concentration in patients with TAA or AAA.
Upon binding to CD4, the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein gp120 undergoes conformational changes that facilitate subsequent interactions with the chemokine coreceptor CXCR4 on the T cells. Our previous study showed that HIV-1 induces breast cancer cell death through gp120-CXCR4 interaction without CD4-induced conformational change of gp120. To characterize the structural properties of CXCR4 on breast cancer cells, the structural differences in CXCR4 between breast cancer cell lines and T cells were investigated. Immunoblots of whole cell lysates from breast cancer cell and T cell lines demonstrated that the predominant forms of CXCR4 on the breast cancer cell lines and T cell lines were three species (45, 61, 100 kDa) and one species (45 kDa), respectively. Cell surface biotin labeling revealed that the 100-kDa polyubiquitinated form of CXCR4 is specifically expressed on the surface of breast cancer cell line DU4475 but not T cell line Molt4#8. The treatment of breast cancer cell lines MDA-MB231 and DU4475 with proteasome inhibitor lactacystin leads to increased surface expression of the 100-kDa polyubiquitinated form of CXCR4 and increases the level of sensitivity to cell death induced by HIV-1. These data suggest that the 100-kDa polyubiquitinated form of CXCR4 plays an important role as a trigger for gp120-induced breast cancer cell death.
Previously, we reported that dibenzylbutyrolactone lignans (DBLLs) from the fruit of Forsythia koreana NAKAI (Oleaceae) has anti-inflammatory, antioxidant, and anti-asthmatic effects. In this study, to clarify the anti-inflammatory mechanisms of DBLL, we evaluated the effects of DBLLs on lipopolysaccharide-stimulated inducible nitric oxide synthetase (iNOS) and cyclooxygenase-2 (COX-2) expressions, nitric oxide (NO) and prostaglandin E2 (PGE2) productions, nuclear factor-κB (NF-κB) and mitogen-activated protein kinase (MAPK) activations, inhibitor of κB (IκB) and inhibitor of κB kinase (IKK) phosphorylations in cytosolic proteins, and cytotoxicity in Raw264.7 cells. DBLLs potently suppressed both the enzyme expression and DNA-binding activity of NF-κB. Arctiin, arctigenin (1.0 μM) and matairesinol (10 μM) inhibited the expression of iNOS by 37.71±2.86%, 32.51±4.28%, and 27.44±2.65%, respectively, and arctiin, arctigenin (0.1 μM) and matairesinol (1.0 μM) inhibited COX-2 expression by 37.93±7.81%, 26.70±4.61% and 29.37±5.21%, respectively. The inhibitory effects of DBLLs on NO and PGE2 productions were the same patterns as those seen for the reductions in iNOS and COX-2 expression, respectively. Arctiin, arctigenin (1.0 μM) and matairesinol (10 μM) significantly (p<0.05) inhibited NF-κB DNA binding by 44.85±6.67%, 44.16±6.61%, and 44.79±5.62%, respectively, and arctiin (0.1 μM) and arctigenin (1.0 μM) significantly (p<0.05) inhibited the phosphorylation of IκB by 20.58±3.86% and 25.99±6.18%, respectively. Furthermore, arctiin, matairesinol (1.0 μM) and arctigenin (10 μM) inhibited the phosphorylation of IKK by 38.80±6.64%, 38.33±6.65%, and 38.57±8.14%, respectively. In addition, DBLLs potently inhibited the lipopolysaccharide (LPS)-induced activation of MAPKs (SAPK/c-Jun NH2-terminal kinase (JNK), p38, and extracellular signal receptor-activated kinase (ERK)1/2). Overall, arctiin was the most effective; its effect was nearly the same as that of 10 μM helenalin. These findings suggest that treatment with non-toxic DBLLs inhibits not only NF-κB and NF-κB-regulated protein activation, but also potently inhibits the activations of specific MAPKs.
In pancreatic islets, free radical formation produced upon exposure to proinflammatory cytokines mediates β cell destruction, which ultimately leads to type 1 diabetes (T1D). In this study, we examined whether laccase, a family of the blue copper protein, can be successfully used to prevent β cells from cytokine-mediated apoptosis. Non-obese diabetic (NOD) mice were used for these experiments. In parallel, the RINm5f β cell line was employed as a model system for in vitro experiments. The results demonstrated that laccase effectively scavenged peroxinitrite, which can be formed by nitric oxide, and upregulated the expression of antioxidant enzymes, such as manganese superoxide dismutase (MnSOD) and catalase. Interestingly, laccase balanced pro- (Bax) and anti-apoptotic (Bcl-2) proteins in terms of both the mRNA and protein levels with a downregulation of cytochrome c protein in RINm5f cells. In addition, laccase maintained blood glucose concentrations at a normal level with a simultaneous increase in plasma insulin levels during the spontaneous induction of diabetes in NOD mice. In conclusion, the antioxidant potentials of laccase in scavenging free radicals and upregulation of antioxidant enzymes may exert its pro-survival effect by counteracting the increased intracellular oxidative stress, and, consequently, by inhibiting apoptosis induced by cytokine-mediated activation during the course of T1D.
Some studies indicated that preventive therapy with Itraconazole oral solution (ITCZ-OS) significantly decreased the incidence of invasive fungal infection, whereas others emphasized that there was no significant decrease. On the other hand, a study involving patients with neutropenia showed a 15-fold increase in the blood concentration of Itraconazole (ITCZ). Therefore, when administering ITCZ-OS, which is more rapidly absorbed in the digestive tract compared to its conventional dosage forms, to patients with blood disease, the blood concentration of ITCZ should be measured to maintain its efficacy and safety. To promote the appropriate use of ITCZ-OS, we conducted blood drug concentration monitoring, and investigated its clinical significance. The subjects were 26 patients with blood diseases. The blood level of ITCZ was measured using HPLC. The mean blood level of ITCZ was 2396.5±1742.7 ng/ml (mean±S.D.). The mean blood level of hydroxy-ITCZ was 5384.4±3348.2 ng/ml. The dose was not correlated with the blood levels of ITCZ/hydroxy-ITCZ per body weight (R2=0.134, 0.154, p=0.094, 0.071). Furthermore, the blood levels of ITCZ and hydroxy-ITCZ per body weight were significantly higher in females (p=0.025, 0.010). In males, there was a correlation between the creatinine clearance and blood level of ITCZ per body weight (R2=0.319, p=0.044). The blood levels of ITCZ varied among the patients. In addition, when ITCZ-OS was administered at a daily dose of 200 mg (ITCZ), the blood levels of ITCZ exceeded a trough level at which this agent may be effective in patients with febrile neutropenia in whom fungal infection is suspected.
The present study examined the influence of cimetidine on the nephrotoxicity and antitumor effects of cisplatin in vitro and in vivo. When the serum concentration of cimetidine was maintained over 20 μg/ml for 4 h by bolus and continuous intravenous infusion, cimetidine prevented nephrotoxicity of cisplatin without influencing antitumor activity. Cimetidine and the antioxidant N-acetylcysteine (NAC) significantly inhibited the in vitro growth inhibition of cisplatin in cells originating from the kidney, but not in SOSN2 osteosarcoma cells. Cimetidine (1 mM) also did not influence platinum concentration in the cells, regardless of whether the organic cation transporter 2 (OCT2) was expressed. Cisplatin did induce reactive oxygen species (ROS) in the KN41 kidney cell line and cimetidine and NAC significantly reduced ROS production. However, cisplatin did not produce ROS in osteosarcoma cells. From these results, cimetidine clearly inhibits nephrotoxicity induced by cisplatin without any influence on the antitumor activity of cisplatin on osteosarcoma in vitro and in vivo.
Some photosensitizing cyanine dyes act on the immune system to enhance the phagocytic capacity of macrophages. In this study, we examined whether these dyes have neurotrophin-like activities and neuroprotective effects in vitro and in vivo. By screening more than 250 cyanine dyes, we found that NK-4 and NK-150, which belong to a group of pentamethine trinuclear cyanine dyes, significantly potentiated nerve growth factor (NGF)-primed neurite outgrowth of PC12HS cells in nanomolar to micromolar concentrations. Both NK-4 and NK-150 showed a remarkable hydroxyl radical-scavenging activity using an in vitro electron spin resonance (ESR)-based technique. They also effectively scavenged peroxy radicals, and in addition, NK-4 acted on superoxides to a similar extent as ascorbate. In vivo, NK-4 and NK-150 prevented cerebral ischemic injury induced by 2 h middle cerebral artery occlusion (MCAO) and 24 h reperfusion in rats. Dyes were intravenously administrated twice 1 h after the occlusion and immediately after the start of reperfusion. NK-4 and NK-150 (100 μg/kg) reduced cerebral infarct volumes by 57.0% and 46.0%, respectively. Those dyes also decreased brain swelling in the ischemic semispheres. As a result, administration of NK-4 and NK-150 provided substantial improvements in MCAO-induced neurological deficits in a dose-dependent manner. These results suggest that NK-4 and NK-150 effectively prevented ischemia-induced brain injury through their potent neurotrophin-like activity as well as antioxidative activity.
We investigated the efficacy of clonazepam as an antiemetic in cisplatin-based chemotherapy for lung cancer. Seven patients experienced cisplatin-induced vomiting despite antiemetic therapy including 5-hydroxytryptamine3 (5-HT3) antagonist and dexamethasone. Therefore, the antiemetic therapy including clonazepam, 5-HT3 antagonist and dexamethasone was subsequently explored in the next course for the same seven patients. We administered clonazepam (0.5 or 1.0 mg/kg) once a day orally for 5 d from day one prior to chemotherapy. The grade of delayed vomiting, evaluated according to Common Terminology Criteria for Adverse Events Version 3.0, in the courses of therapy with clonazepam was significantly lower than without clonazepam (p=0.013). The patients whose serum clonazepam concentrations were below the lower limit of detection (3.0 ng/ml) experienced vomiting in three of three courses, whereas the patients whose serum clonazepam concentrations were higher than 4.3 ng/ml experienced no vomiting in six of seven courses. We observed that the symptom of cisplatin-induced delayed vomiting is controlled with serum clonazepam levels in the order of 10.0 ng/ml.
A homology-based cloning strategy yielded two cDNA clones designated Am-cam-1 and Am-cam-2, presumably encoding calmodulin protein from a callus culture derived from the leaf tissues of Aquilaria microcarpa. An appreciable increase in the transcriptional activity of Am-cam-1 was reproducibly observed by exposure of the cell culture to methyl jasmonate, as analyzed by a reverse-transcription polymerase chain reaction. The expression level of the gene also increased when the cells were treated with yeast extract. The transcription of Am-cam-2 was similarly stimulated by the treatment with methyl jasmonate and yeast extract, however, the intensities of the enhanced expression appeared to be lower as compared with that of Am-cam-1. In contrast, Ca2+-ionophore A23187 did not show inducing activity for the expression of these two calmodulin genes. These results suggest that Am-cam-1 and Am-cam-2 and their products play important roles in signal transduction processes in methyl jasmonate- and yeast extract-treated cells of A. microcarpa, accompanying the change in the transcriptional activities.
Antigen-presenting cells are key vehicles for delivering antigens in tumor immunotherapy, and the most potent of them are dendritic cells (DCs). Recent studies have demonstrated the usefulness of DCs genetically modified by lipofection in tumor immune therapy, although sufficient gene transduction into DCs is quite difficult. Here, we show that Paeoniae radix, herbal medicine, and the constituent, 1,2,3,4,6-penta-O-galloyl-β-D-glucose (PGG), have an attractive function to enhance phagocytosis in murine dendritic cell lines, DC2.4 cells. In particular, PGG in combination with lipofectin (LPF) enhanced phagocytic activity. Furthermore, PGG enhanced lipofection efficacy in DC2.4 cells, but not in colorectal carcinoma cell lines, Colon26. In other words, PGG synergistically enhanced the effect of lipofectin-dependent phagocytosis on phagocytic cells. Hence, according to our data, PGG could be an effective aid in lipofection using dendritic cells. Furthermore, these findings provide an expectation that constituents from herbal plant enhance lipofection efficacy.
The Renkin function was applied to characterize the penetration pathways through rat skin following different pretreatments. Nonmetabolic oligosaccharides and sugar alcohols, as model hydrophilic compounds, were applied simultaneously to the excised skin to obtain the equivalent cylindrical pore radius (R) and pore occupancy/length ratio (ε/L) for each skin piece. The R and ε/L values obtained were used to construct the simulation curves of the permeability coefficient (Pa)–molecular weight (MW). In the case of full-stripped skin, the Pa of the model compounds and separately obtained Pa of 5(6)-carboxyfluorescein (CF) showed good agreement with the simulation curve based on the Renkin function, suggesting that the viable epidermis and dermis in the full-stripped skin contained permeation pathways for hydrophilic compounds like aqueous channels. On the other hand, there was poor agreement of Pa with the simulation curve for skin pretreated with an ethanol-menthol mixed enhancer system and the observed Pa of CF in the pretreated skin was twice that calculated. The enhancer system might not be able to create aqueous channels in the lipid layer of the stratum corneum and could increase the permeation of CF in the layer in a different way. The analysis presented here will be useful not only for quantitative evaluation of drug permeation through aqueous channels in treated skins but also for investigation of the mechanism of skin-permeation enhancing techniques.
Medicinal pteridophytes are an important group used in traditional Chinese medicine; however, there is no simple and universal way to differentiate various species of this group by morphological traits. A novel technology termed “DNA barcoding” could discriminate species by a standard DNA sequence with universal primers and sufficient variation. To determine whether DNA barcoding would be effective for differentiating pteridophyte species, we first analyzed five DNA sequence markers (psbA-trnH intergenic region, rbcL, rpoB, rpoC1, and matK) using six chloroplast genomic sequences from GeneBank and found psbA-trnH intergenic region the best candidate for availability of universal primers. Next, we amplified the psbA-trnH region from 79 samples of medicinal pteridophyte plants. These samples represented 51 species from 24 families, including all the authentic pteridophyte species listed in the Chinese pharmacopoeia (2005 version) and some commonly used adulterants. We found that the sequence of the psbA-trnH intergenic region can be determined with both high polymerase chain reaction (PCR) amplification efficiency (94.1%) and high direct sequencing success rate (81.3%). Combined with GeneBank data (54 species cross 12 pteridophyte families), species discriminative power analysis showed that 90.2% of species could be separated/identified successfully by the TaxonGap method in conjunction with the Basic Local Alignment Search Tool 1 (BLAST1) method. The TaxonGap method results further showed that, for 37 out of 39 separable species with at least two samples each, between-species variation was higher than the relevant within-species variation. Thus, the psbA-trnH intergenic region is a suitable DNA marker for species identification in medicinal pteridophytes.