Egg-Hatching Mechanism of Human Liver Fluke, Opisthorchis viverrini: A Role For Leucine Aminopeptidases From the Snail Host, Bithynia siamensis goniomphalos.

The human liver fluke Opisthorchis viverrini (Platyhelminthes, Trematoda, Digenea) uses snails of the genus Bithynia as first intermediate host. Peculiarly among trematodes, the eggs of O. viverrini hatch within the digestive tract of its snail host. It remains uncertain whether hatching in this species is mediated through mechanical fracture of the eggshell or by digestion with specific digestive enzymes. This study aimed to characterize enzymes with specific inhibitors and factors involved in the hatching activity of O. viverrini eggs. For measuring egg hatching in vivo, 50 O. viverrini mature eggs were fed to individual Bithynia siamensis goniomphalos snails at various temperature conditions for 24 hr. Ex vivo, mature eggs were incubated with crude snail extract and commercial leucine aminopeptidase (LAP). Egg-hatching of O. viverrini was temperature dependent, with optimal hatching occurring at 24-28 C, with a peak of hatching of 93.54% in vivo and 30.55% ex vivo occurring at these temperatures. Ex vivo hatching rates increased to 45.87% under anaerobic conditions at 28 C. Some 22.70% and 16.21% of heat-killed eggs also hatched within the snail digestive tract and snail extract, respectively, indicating that host molecules are involved in the hatching response. Most eggs hatch in the anterior regions of the digestive tract. Hatching was completely inhibited in the presence of bestatin, an inhibitor of LAP, but not in the presence of phosphatase inhibitors. Bestatin inhibition of hatching was reversible. Finally, egg hatching could be induced by addition of a porcine LAP. The results indicate that this digenean utilizes both LAP of the snail host and movement of miracidia for hatching.

A phase II randomized double-blind placebo-controlled study of 6-gingerol as an anti-emetic in solid tumor patients receiving moderately to highly emetogenic chemotherapy.

6-Gingerol is a natural compound extracted from ginger. Preclinical studies demonstrated that 6-gingerol has an anti-emetic activity by inhibiting neurokinin-1, serotonin, and dopamine receptors. Several clinical trials examined crude ginger powder for preventing chemotherapy-induced nausea and vomiting (CINV), but none of them was conducted with a standardized bioactive compound. Patients who received moderately to highly emetogenic adjuvant chemotherapy were randomized to receive 6-gingerol 10 mg or placebo orally twice daily for 12 weeks. Ondansetron, metoclopramide, and dexamethasone were given to all patients. The primary endpoint was complete response (CR) rate defined as no emesis or rescue treatment at any time. Eighty-eight patients were randomized to receive 6-gingerol (N = 42) or placebo (N = 46). Most patients received highly emetogenic chemotherapy (93%). Overall CR rate was significantly higher in 6-gingerol group as compared with that of the placebo (77 vs. 32%; P < 0.001). The difference in means of appetite score was significant (P = 0.001) and more noticeable over time. Mean FACT-G score indicating quality of life was significantly higher (86.21) in 6-gingerol group at 64 days as compared with that of placebo group (72.36) (P < 0.001). No toxicity related to 6-gingerol was observed. Patients treated with 6-gingerol reported significantly less grade 3 fatigue (2 vs. 20%; P = 0.020). 6-Gingerol significantly improved overall CR rate in CINV, appetite and quality of life in cancer patients receiving adjuvant chemotherapy. A phase III randomized study of 6-gingerol is warranted to confirm these results.

Modulation of P-glycoprotein function and multidrug resistance in cancer cells by Thai plant extracts.

The effects of ethanol extracts from Thai plants belonging to the families of Annonaceae, Rutaceae, and Zingiberaceae on P-glycoprotein (P-gp) function and multidrug resistance were examined in paclitaxel-resistant HepG2 (PR-HepG2) cells. All the extracts tested, significantly increased the accumulation of [3H]paclitaxel, a P-gp substrate, in the cells. Among nine extracts, Z01 and Z02, extracts from Curcuma comosa and Kaempferia marginata (Zingiberaceae family), respectively, potently increased the accumulation. In addition, Z01 and Z02 increased the accumulation of other P-gp substrates, rhodamine 123 and doxorubicin, in PR-HepG2 cells in a concentration-dependent manner. Increased accumulation of rhodamine 123 and doxorubicin by Z01 and Z02 was also confirmed by confocal laser scanning microscopy. The effect of Z01 and Z02 pretreatment on the expression of MDR1 mRNA was also examined. The expression of MDR1 mRNA was not affected by the treatment of PR-HepG2 cells with these extracts for 48 hours. Cytotoxicity of paclitaxel was examined by XTT and protein assays in the absence and presence of Z02. Z02 potentiated the cytotoxicity of paclitaxel in PR-HepG2 cells. These results suggest that Curcuma comosa and Kaempferia marginata belonging to Zingiberaceae are useful sources to search for new P-gp modulator(s) that can be used to overcome multidrug resistance of cancer cells.

A prospective, randomized clinical trial comparing topical aloe vera with 0.1% triamcinolone acetonide in mild to moderate plaque psoriasis.

BACKGROUND: Topical aloe vera (AV) has been used to treat various skin conditions, including psoriasis, with good results. OBJECTIVES: This study aims to compare the efficacy of AV and 0.1% triamcinolone acetonide (TA) in mild to moderate plaque psoriasis. METHODS: A randomized, comparative, double-blind, 8-week study was designed. Eighty patients randomly received AV or 0.1% TA cream and their clinical response were evaluated using the Psoriasis Area Severity Index (PASI) and the Dermatology Life Quality Index (DLQI). RESULTS: After 8 weeks of treatment, the mean PASI score decreased from 11.6 to 3.9 (-7.7) in the AV group and from 10.9 to 4.3 (-6.6) in the TA group. Between-group difference was 1.1 (95% confidence interval -2.13, -0.16, P = 0.0237). The mean DLQI score decreased from 8.6 to 2.5 (-6.1) in the AV group and from 8.1 to 2.3 (-5.8) in the TA group. Between-group difference was 0.3 (95% confidence interval -1.18, -0.64, P = 0.5497). There was no follow-up period after the 8-week treatment. CONCLUSIONS: AV cream may be more effective than 0.1% TA cream in reducing the clinical symptoms of psoriasis; however, both treatments have similar efficacy in improving the quality of life of patients with mild to moderate psoriasis.

The efficacy of aloe vera gel in the treatment of oral lichen planus: a randomized controlled trial.

BACKGROUND: Oral lichen planus (OLP) is a chronic inflammatory disease that can be painful especially in the atrophic and erosive forms. Several therapies have been tried, with varying results. There is one case report in which aloe vera (AV) was used successfully in the treatment of lichen planus. OBJECTIVES: To compare the efficacy of AV and placebo in the topical management of OLP. METHODS: A randomized, double-blind, placebo-controlled trial was designed. Fifty-four patients were randomized into two groups to receive AV gel or placebo for 8 weeks. RESULTS: Fifty-four consecutive patients (34 women and 20 men) participated in the study. We found erosive and ulcerative lesions in 83% and 17%, respectively. The most common site of OLP was the lower lip. Twenty-two of 27 patients treated with AV (81%) had a good response after 8 weeks of treatment, while one of 27 placebo-treated patients (4%) had a similar response (P<0.001). Furthermore, two patients treated with AV (7%) had a complete clinical remission. Burning pain completely disappeared in nine patients treated with AV (33%) and in one treated with placebo (4%) (P=0.005). Symptomatology improved by at least 50% (good response) in 17 patients treated with AV (63%) and in two treated with placebo (7%) (P<0.001). No serious side-effects were found in both groups. CONCLUSIONS: AV gel is statistically significantly more effective than placebo in inducing clinical and symptomatological improvement of OLP. Therefore, AV gel can be considered a safe alternative treatment for patients with OLP.

Simultaneous identification and quantitation of 11 flavonoid constituents in Kaempferia parviflora by gas chromatography.

Kaempferia parviflora (Krachaidum; KD) is a Thai herb, the rhizomes of which have been used in folk medicine and ritual ceremonies. The increasing use of KD has led to concerns regarding the variation of quality, potency and efficacy of KD products. A gas chromatographic method was developed and validated using 11 flavonoids that had been fully characterized as reference. Limits of detection ranged from a low of 0.1 ppm to a high of 1.0 ppm. The limits of quantitation were a low of 0.5 ppm (5-hydroxy-3,7-dimethoxyflavone) to a high of 3.0 ppm (5,7,4'-trimethoxyflavone and 5,7,3',4'-tetramethoxyflavone). Precision of intra- and inter-day analyses gave a RSD range of 3.02-8.25 and 2.84-12.37, respectively. The diversity of flavonoid content and their distribution profiles in KD samples from 12 different origins was investigated using the validated method. Total flavonoid content in these samples ranged from 23.86 to 60.98 mg/g. Two of the compounds, 5,7-dimethoxyflavone and 5,7,4'-trimethoxyflavone, emerged as major constituents. Samples contained as much as 21.68 and 9.88 mg/g, respectively. Two distinct patterns of the distribution of the flavonoids, as characterized by the ratio of these two compounds in the KD rhizome samples, were observed. This method is expected to be useful in the quantitative and qualitative analyses of the flavonoid content of KD samples and as a quality control assessment of KD raw materials and products.

Antimutagenic and anticarcinogenic effects of Phyllanthus amarus.

This study aimed to examine the antimutagenic and anticarcinogenic potential of Phyllanthus amarus Schum. et Thonn. using the bacterial preincubation mutation assay and an in-vivo alkaline elution method for DNA single-strand breaks in hamster liver cells. The aqueous extract of the entire plant showed an antimutagenic effect against induction by 2-aminofluorene (AF2), 2-aminoanthracene (2AA) and 4-nitroquinolone-1-oxide (4-NQO) in Salmonella typhimurium strains TA98 and TA100, and in Escherichia coli WP2 uvrA/pKM101. All the results were dose-dependent; however, inhibition of N-ethyl-N-nitrosoguanidine (ENNG)-induced mutagenesis was observed only with S. typhimurium TA100. The extract also exhibited activity against 2-nitrofluorene (2NF) and sodium azide-induced mutagenesis with S. typhimurium TA98 and TA100, respectively. Based on the alkaline elution method, the plant extract prevented in vivo DNA single-strand breaks caused by dimethylnitrosamine (DMN) in hamster liver cells. When the extract was administered 30 min prior to the administration of DMN, the elution rate constant decreased more than 2.5 times, compared to that of control. These results indicate that P. amarus possesses antimutagenic and antigenotoxic properties.

Diuretic effects of selected Thai indigenous medicinal plants in rats.

Extracts of five indigenous Thai medicinal having ethnomedical application in the treatment of dysuria were investigated for their diuretic activity. Root extracts of Ananas comosus and Carica papaya, given orally to rats at a dose of 10 mg/kg, demonstrated significantly increased urine output (P < 0.01) which was 79 and 74%, respectively, of the effect of an equivalent dose of hydrochlorothiazide. Both plant extracts gave similar profiles of urinary electrolyte excretion to that of the hydrochlorothiazide. The analyses of the urinary osmolality and electrolyte excretion per unit time suggest the observed effect of A. comosus was intrinsic, whereas that of C. papaya may have resulted from a high salt content of this extract. However, our experimental evidence on the diuretic activities of the other three plants did not parallel their local utilization for dysuria. It was found that the rhizome of Imperata cylindrica apparently inhibited the urination of rats whereas the rhizome of Cyperus rotundus and the stem of Averrhoa carambola failed to demonstrate any diuretic activities. These results indicate that two of the plants investigated exert their action by causing diuresis. The other three plants need further investigation to determine their effectiveness in the treatment of dysuria.

Two renal alpha 2-adrenergic receptor sites revealed by p-aminoclonidine binding.

[3H]p-aminoclonidine [3H]PAC, a specific alpha 2-adrenergic agonist, was used to characterize alpha 2-adrenoceptor binding in rat renal membranes. Rosenthal plots demonstrated two binding sites with Kds of approximately 1.7 and 14.2 nM and Bmaxs (maximum binding) of 47.3 and 218.8 fmol/mg protein for the high- and low-affinity sites, respectively. These characteristics were confirmed by estimate of Kd parameters based on association and dissociation experiments. Pseudo-Hill coefficients generated from drug inhibition experiments were all less than unity, suggesting differential binding at two alpha 2-adrenoceptor binding sites. Specific alpha 2-adrenergic agonists exhibited greater binding affinity to both sites than did nonspecific drugs, and all drugs displayed greater affinity for the high- than the low-affinity binding site. Both guanyl nucleotides and sodium chloride inhibited [3H]PAC binding more at the high-affinity than at the low-affinity site. Renal denervation resulted in significant upregulation of receptor density only at the high-affinity sites with no change in receptor affinity at either site, suggesting that a majority of the alpha 2-adrenoceptors in the kidney are postsynaptic. Thus all lines of evidence in this study indicate that two alpha 2-adrenoceptor binding sites exist in the rat kidney.

The development of alpha 2-adrenoceptors in the rat kidney: correlation with noradrenergic innervation.

The development of the renal alpha 2-adrenoceptors in the rat was studied by binding renal membranes of prenatal and postnatal rats with p-[3,5-3H]aminoclonidine (PAC), a selective alpha 2-adrenoceptor agonist. The results demonstrate that prior to birth and on the day of birth [3H]PAC binding is present and reflects a single linear binding affinity (Kd approximately equal to 1.0 nM). In contrast, data from postnatal day 3 demonstrate the addition of a second, lower affinity binding site (Kd approximately equal to 7.0 nM) that develops rapidly during the ensuing weeks of life. Both high and low affinity alpha 2-adrenoceptor binding sites continue to increase in receptor density through 30 weeks of age; however, the binding affinity of each site remains relatively stable. The sympathetic noradrenergic innervation of the rat kidney is present prenatally and rapidly develops during the first 3 weeks of life with peak noradrenaline concentrations being reached at approximately prenatally and rapidly develops during the first 3 weeks of life with peak noradrenaline concentrations being reached at approximately postnatal day 21. Immunohistochemical staining demonstrates that tyrosine hydroxylase-positive axons rapidly develop in the kidney during the same period, indicating that the increase in noradrenaline is related to expansion of renal sympathetic innervation.

An autoradiographic analysis of the time of origin of neurons in the hypothalamus of the cat.

The pattern of neurogenesis of the cat hypothalamus was studied by the use of the [3H]thymidine autoradiographic method. The neurons of the cat hypothalamus are nearly all generated in a period from embryonic days E21 to E35, and in most cases the neurons in a single nucleus are generated over a much shorter period. The dominant gradient along which the cells are organized is lateral-medial (outside-in). This gradient was apparent in all nuclei except the periventricular and arcuate nuclei. Two other gradients were observed in some, but not all, nuclei. These were along the dorsoventral and the posteroanterior axis. The dorsoventral gradient was uniformly less pronounced than the lateromedial gradient. The posteroanterior gradient was most obvious in the mammillary complex. An analysis of the data suggests that the neurons of individual nuclei do not necessarily have a unique production history. This suggests that insofar as nuclear formation is concerned, factors such as the parental population generating the neurons that populate a nucleus, as well as the early neuronal interconnections, may play a more important role than the birthdates themselves.

The development of Ammon's horn and the fascia dentata in the cat: a [3H]thymidine analysis.

The present [3H]thymidine autoradiographic analysis of neurogenesis demonstrates that the neurons which populate the adult cat hippocampus are born between embryonic day (E)22 and E42. In contrast, although neuronal production in the fascia dentata begins on the same day, granule cells in this area continue to be produced throughout prenatal life and into early postnatal life, and probably continues at an extremely low rate well into adulthood. Three major sets of spatiotemporal gradients characterize the production of neurons in Ammon's horn and the fascia dentata. The first set involves the radial axis. Within the hippocampus there exists an inside-out gradient. The reverse gradient is present in the fascia dentata, i.e. outside-in. The second set of gradients involves the transverse or rhinodentate axis. In general the CA3 neurons are born earlier than the CA1 neurons. Within both neuronal layers of the fascia dentata, the hidden blade cells tend to be born earlier than those of the exposed blade. Again, the pattern in the fascia is the reverse of that in the hippocampus proper. A temporal to septal gradient is also present, but this is the weakest of the gradients.

The cortical projection of the basolateral amygdaloid nucleus in the rat: a retrograde fluorescent dye study.

The fluorescent dye, retrograde labeling technique was used to determine the extent of the projection from the basolateral nucleus of the amygdala to the neocortex in the rat. Each rat received a single cortical injection of fast blue, and in one-half of the animals, a subsequent injection of nuclear yellow was placed in a different cortical region. An analysis of the results demonstrates that the projection to the midline cortex arises in the medial neurons within the caudal two-thirds of the basolateral nucleus. This projection is directed to the anterior cingulate cortex, but not to the posterior cingulate cortex. The primary motor cortex receives a basolateral amygdala projection which originates from neurons in two areas, (1) the medial part of the anterior one-third of the nucleus and (2) the center (in the lateral to medial axis) portion of the posterior two-thirds of the nucleus. The latter neurons are situated lateral to the neurons projecting to the cingulate cortex. Somatosensory cortex injections label many fewer basolateral nucleus neurons than do motor cortex injections, but these neurons are located in a position similar to that of those labeled by motor cortex injections. Finally the gustatory cortex, which lies just dorsal to the rhinal sulcus, receives a basolateral projection from neurons in the lateroventral one-half of the basolateral nucleus. These results demonstrate that the basolateral nucleus gives rise to a rather widespread and topographically organized projection to the anterior half of the neocortex of the rat.

An analysis of the time of origin of neurons in the entorhinal and subicular cortices of the cat.

The [3H]thymidine autoradiographic method was used to determine the birth dates of neurons in the cat parahippocampal gyrus. Cat fetuses were exposed to a single pulse of the radioactive marker between the 20th and 55th embryonic days. All animals were delivered normally and allowed to survive for 2-6 months postnatal. The resulting autoradiographs demonstrate three spatiotemporal gradients of cell birth in the entorhinal and subicular cortices. First, an inside-out gradient is apparent; i.e., neurons in the deeper layers are born earlier than those in the more superficial layers. Second, a rhino to dentate gradient exists. Accordingly, cells closer to the lateral entorhinal region tend to be generated earlier than those further away. Third, a temporal to septal gradient is present. Neurons close to the anterior pole of the temporal lobe are born earlier than those more caudally located. Whereas the first two gradients have been observed in other species, the latter gradient has not been reported consistently. Three exceptions to these overall gradients exist. First, neurons near the layer I/II border are born earlier than the majority of the layer II neurons. Second, neurons near the transition zone between two adjacent regions are born earlier than neurons located in the middle of each region. Third, the prosubiculum and subiculum do not exhibit a clear inside-out or temporal to septal gradient.