The acute toxicity of water hemlock (Cicuta douglasii) in a goat model.

Water hemlock (Cicuta douglasii) is one of the most toxic plants to livestock and humans. Little is known regarding the amount of plant required to cause death. The objective of this study was to determine a lethal dose of water hemlock in a goat model. Plants were dosed to goats via oral gavage of freeze-dried ground plant material. The results from this study suggest that 1-2 fresh tubers would be lethal to goats.

White snakeroot poisoning in goats: Variations in toxicity with different plant chemotypes.

Tremetone and possibly other benzofuran ketones are believed to be the toxic compounds in white snakeroot. However, disease has not been reproduced with purified toxins and the concentrations of the benzofuran ketones in white snakeroot populations that cause toxicosis have not been documented. The objectives of this study were to compare the toxicity of seven plant populations, better characterize the clinical and pathologic changes of poisoning, and correlate intoxication with benzofuran ketone content. Four of the seven white snakeroot collections were toxic at the dose and duration used in the study. Affected goats became exercise intolerant, had significant serum enzyme changes and histological lesions in the large appendicular muscles. The incidence and severity of poisoning was not correlated with total doses of tremetone or total benzofuran ketone concentrations suggesting they may not be closely involved in producing toxicity and the possible involvement of an unidentified toxin. The results also demonstrate that white snakeroot populations vary chemically and toxicologically.

An open-label trial of Korean red ginseng as an adjuvant treatment for cognitive impairment in patients with Alzheimer's disease.

BACKGROUND AND PURPOSE: Ginseng is one of the most popular herbs worldwide. Ginseng has various medical applications, and it seems to have significant effects as a cognition-enhancing drug. In this study, we examined the efficacy of Korean red ginseng (KRG) as an adjuvant therapy to conventional anti-dementia medications in patients with Alzheimer's disease. METHODS: The trial was designed as a 12-week randomized study. Sixty-one patients (24 males and 37 females) with Alzheimer's disease were randomly assigned to one of the following treatment groups: low-dose KRG (4.5 g/day, n = 15), high-dose KRG (9 g/day, n = 15) or control (n = 31). The Alzheimer's Disease Assessment Scale (ADAS), Korean version of the Mini-Mental Status Examination (K-MMSE) and Clinical Dementia Rating (CDR) scale were used to assess the change in cognitive and functional performance at the end of the 12-week study period. RESULTS: The patients in the high-dose KRG group showed significant improvement on the ADAS and CDR after 12 weeks of KRG therapy when compared with those in the control group (P = 0.032 and 0.006 respectively). The KRG treatment groups showed improvement from baseline MMSE when compared with the control group (1.42 vs. -0.48), but this improvement was not statistically significant. CONCLUSIONS: KRG showed good efficacy for the treatment of Alzheimer's disease; however, further studies with larger samples of patients and a longer efficacy trial should be conducted to confirm the efficacy of KRG.

Evaluation of vaccination against methyllycaconitine toxicity in mice.

The purpose of this study was to determine whether larkspur toxins conjugated to protein carriers would promote active immunity in mice. Mice were injected with several larkspur toxin-protein conjugates or adjuvant alone to determine whether the resulting immunological response altered animal susceptibility to methyllycaconitine, the major toxic larkspur alkaloid. Although vaccinations increased the calculated lethal dose 50% (LD50) for intravenous methyllycaconitine toxicity, overlapping confidence intervals did not provide evidence of differences between the vaccinated and control groups. In the lycoctonine conjugate (LYC)-vaccinated group, mouse survival was related (P = 0.001) to serum titers for methyllycaconitine doses up to 4.5 mg/kg of body weight. When mice withlow antibody titers were removed from the vaccinated groups in which titer was related to survival, the recalculated LD50 estimates were 20% greater than the LD50 of the control group. However, the 95% confidence intervals of the recalculated LD50 groups overlapped with the control groups. Overall, these results suggest that vaccination altered methyllycaconitine toxicity in mice and that vaccination may be useful in decreasing the effects of larkspur toxins in animals. Additional studies are warranted to continue development of potential larkspur vaccines for livestock.

Protective effects of curcumin against oxidative damage on skin cells in vitro: its implication for wound healing.

BACKGROUND: Curcumin, isolated from turmeric, has been known to possess many pharmacologic properties. It has been proven to exhibit remarkable anticarcinogenic, anti-inflammatory, and antioxidant properties. Turmeric curcumin may be a good potential agent for wound healing. METHODS: To further understand its therapeutic mechanisms on wound healing, the antioxidant effects of curcumin on hydrogen peroxide (H2O2) and hypoxanthine-xanthine oxidase induced damage to cultured human keratinocytes and fibroblasts were investigated. Cell viability was assessed by colorimetric assay and quantification of lactate dehydrogenase release. RESULTS: Exposure of human keratinocytes to curcumin at 10 microg/mL showed significant protective effect against hydrogen peroxide. Interestingly, exposure of human dermal fibroblasts to curcumin at 2.5 microg/mL showed significant protective effects against hydrogen peroxide. No protective effects of curcumin on either fibroblasts or keratinocytes against hypoxanthine-xanthine oxidase induced damage were found in our present studies. CONCLUSION: The findings indicate that curcumin indeed possessed powerful inhibition against hydrogen peroxide damage in human keratinocytes and fibroblasts.

Evaluating the role of alternative therapy in burn wound management: randomized trial comparing moist exposed burn ointment with conventional methods in the management of patients with second-degree burns.

CONTEXT: Moist exposed burn ointment (MEBO), from China, has been said to revolutionize burn management. OBJECTIVE: Our study was conducted to compare MEBO with conventional management (C) with respect to the rate of wound healing, antibacterial and analgesic effect, and hospital costs. DESIGN: This is a prospective, randomized, controlled clinical trial conducted between 1 March 1997 and 24 October 1998. SETTING: The trial was conducted in a specialized burn facility located in a tertiary referral hospital in a developed and industrialized island-state in Southeast Asia. PATIENTS: We randomly assigned 115 consecutive patients between the ages of 12 and 80 who had partial-thickness thermal burns covering less than 40% of body surface area (BSA) to receive either MEBO or C. Fifty-seven patients were assigned to MEBO and 58 patients to C. The latter group received twice-daily dressing changes; MEBO patients received MEBO every 4 hours. MAIN OUTCOME MEASURES: Patients were hospitalized until 75% BSA had healed. BSA was determined by visual inspection and charted on Lund and Browder charts regularly. Wound healing rate, bacterial infection rate, pain score, and hospitalization costs were recorded. RESULTS: The median time to 75% healing was 17.0 and 20.0 days with MEBO and C, respectively (HR = 0.67, 95% CI = 0.41-1.11, P =.11), suggesting similar efficacy between the 2 modalities. Bacterial infection rates were similar between the 2 groups (HR = 1.10, 95% CI = 0.59-2.03, P =.76). MEBO imparted a greater analgesic effect in the first 5 days of therapy and reduced hospital costs by 8%. CONCLUSIONS: MEBO is as effective as conventional management but is not the panacea for all burn wounds. The use of MEBO eases the management of face and neck burns and facilitates early institution of occupational therapy in hand burns. It confers better pain relief such that fewer opiates are used during the first 5 days after burn injury.

Development of enzyme-linked immunosorbent assays for the hepatotoxic alkaloids riddelliine and riddelliine N-oxide.

Pyrrolizidine alkaloid-containing plants are widely distributed throughout the world and are particularly common in the genus Senecio. The structural types and concentrations of the alkaloids vary among plant species. In addition, within a species of plant, concentrations vary with environment and location. Many pyrrolizidine alkaloids are toxic and cause poisoning in livestock and in humans. Rapid, sensitive, and specific diagnostic techniques are needed to identify poisoned animals and to determine the particular plants and conditions under which livestock are likely to be poisoned. In this study, two competitive inhibition enzyme-linked immunosorbent assays for riddelliine, riddelliine N-oxide, and other closely related pyrrolizidine alkaloids were developed using polyclonal antibodies. One assay is class specific toward the free base forms of the pyrrolizidine alkaloids; the other assay showed cross-reactivity to both the free base and N-oxide forms of the alkaloids. The assay with the lowest limit of detection had an I(50) of 803.9 pg with a limit of detection of 47.5 pg for riddelliine. Spike and recovery studies for riddelliine in bovine blood ranged from 45 to 74%. The assay that showed cross-reactivity between the N-oxide and free base forms of the pyrrolizidine alkaloids allowed estimation of the total pyrrolizidine alkaloid content in Senecio riddellii in admixture with alfalfa. These findings suggest that these techniques will be excellent tools to diagnose poisoned animals and identify highly toxic plants.

Phenolic compounds of Chromolaena odorata protect cultured skin cells from oxidative damage: implication for cutaneous wound healing.

Extracts from the leaves of Chromolaena odorata have been shown to be beneficial for treatment of wounds. The crude ethanol extract of the plant had been demonstrated to be a powerful antioxidant to protect fibroblasts and keratinocytes in vitro. In this study, the most active compounds were fractionated and identified from the crude extract using liquid chromatography coupled with UV spectroscopy and mass spectrometry. The antioxidant effects of purified fractions on cultured fibroblasts and keratinocytes were investigated using colorimetric and lactate hydrogenase release assay. The results showed that the phenolic acids present (protocatechuic, p-hydroxybenzoic, p-coumaric, ferulic and vanillic acids) and complex mixtures of lipophilic flavonoid aglycones (flavanones, flavonols, flavones and chalcones) were major and powerful antioxidants to protect cultured skin cells against oxidative damage. In conclusion, the extract from C odorata contains a mixture of powerful antioxidant compounds that may be one of potential mechanism contributing to enhanced wound healing.

Development of enzyme-linked immunosorbent assays for toxic larkspur (Delphinium spp.) alkaloids.

Larkspur (Delphinium spp.) poisons thousands of cattle on western rangelands each year. Because poisoning does not cause specific lesions, and poisoned animals are rarely found before they die, definitively identifying poisoned animals is difficult. Additionally, toxin concentrations in larkspur plants vary with environment, plant, and location. Rapid, sensitive, and specific diagnostic techniques are needed to identify poisoned animals and to determine when and what plants are likely to poison livestock. In this study, three competitive inhibition enzyme-linked immunosorbent assays (CI-ELISA) for toxic larkspur alkaloids were developed. One assay is class-specific toward the N-(methylsuccinimido)anthranoyllycoctonine (MSAL) alkaloids, and two assays are specific for individual alkaloids. The assay with the lowest limit of detection had an I(50) of 191 pg with a limit of detection of 30.5 pg for methyllycaconitine. Spike and recovery studies using bovine blood and brain tissue ranged from 52 to 89%. These findings suggest that with additional development these techniques are likely to be excellent tools for diagnosing poisoned animals and identifying highly toxic plants.

The role of alternative therapy in the management of partial thickness burns of the face--experience with the use of moist exposed burn ointment (MEBO) compared with silver sulphadiazine.

INTRODUCTION: Conventional management of partial thickness facial burn wounds includes the use of silver sulphadiazine dressings. Silver sulphadiazine forms an overlying slough that makes wound healing assessment difficult. Moist exposed burn ointment (MEBO) has been proposed as the ideal burn wound dressing both for burns of the face and other sites. Proponents of MEBO claim that it accelerates wound healing and results in scarless wound healing and at the same time reduce bacterial colonisation and the need for analgesics. We present here our experience with MEBO in the management of partial thickness burns of the face. MATERIALS AND METHODS: One hundred and fifteen patients with partial thickness burns were randomly assigned to conventional treatment or MEBO. Out of this, 112 were analysed. Thirty-nine patients sustained facial burns; 17 received MEBO and 22 received silver sulphadiazine. Patients were followed up daily until the burn wounds were reduced by 75% of original body surface area (BSA). RESULTS: In patients with facial burns, MEBO was similar to silver sulphadiazine therapy with respect to rate of wound healing. Minimal slough was present over the wounds in MEBO-treated wounds resulting in clearer assessment of healing progression. CONCLUSIONS: Advantages of MEBO as compared to silver sulphadiazine in the management of partial thickness burns of the face include convenient change of dressing and easier assessment of healing progression. This suggests that MEBO is a useful alternative therapy for partial thickness burns of the face.

Investigating plant-based medicines for wound healing with the use of cell culture technologies and in vitro models: a review.

INTRODUCTION: Cell culture and molecular technologies are basic yet sophisticated research tool used to investigate plant-based medicine for wound healing. METHODS: Cell viability and proliferation assay is used to determine whether there are any positive effects and to discover what is the limiting cytotoxic concentration in vitro. The scratch technique, fibroblast-populated collagen lattices and aortic rings embedded gels are used as the in-vitro models of wound re-epithelialization, contraction and angiogenesis. The immunofluorescence, immunoblotting and organotypic culture can be used to detect expression of specific proteins that are modulated by plant extracts during the wound healing process. MAIN FINDINGS: Given the dynamics of the wound healing process, cell culture and molecular technologies are advantageous in providing us with detailed studies and analysis of each intricate process. CONCLUSION: The scientific approaches for the study of traditional plant-based remedies for wound healing will provide us an important platform for rigorous testing and evaluation of their clinical efficacy based on accepted rules of evidence.

Exon-intron structure of the human PTK6 gene demonstrates that PTK6 constitutes a distinct family of non-receptor tyrosine kinase.

Partial PTK6 (also known as Brk) cDNA was initially isolated by reverse transcription-PCR of normal human melanocyte mRNAs and the full-length cDNA encodes a non-receptor protein tyrosine kinase with an SH3 domain, an SH2 domain, and a kinase catalytic domain. We have cloned the human PTK6 gene by screening human genomic lambda libraries using the full-length PTK6 cDNA as probe. The human PTK6 gene consists of 8 exons encompassing 8.8 kb and all the splicing junctions followed the conserved GT/AG rule. Coding sequence of the PTK6 gene was identical to that of the cDNA cloned from T-47D, human breast tumor cell line. Although the amino acid sequence of the PTK6 polypeptide showed the strongest homology to those of the Src family members of protein tyrosine kinases, exon-intron boundaries of the PTK6 gene were quite different from those of the Src family genes, which are evolutionarily conserved. The 813-bp 5'-flanking sequence of the PTK6 gene upstream of a luciferase reporter gene conferred significant promoter activity, at approximately 60% level of the SV40 promoter, in transient expression assays into MCF-7, human breast tumor cell line. PTK6 mRNA was expressed at very high level in colon and at high levels in small intestine and prostate, and at low levels in some tested fetal tissues. These results suggest that PTK6 constitutes an evolutionarily distinct family of non-receptor protein tyrosine kinases and may function as an intracellular signal transducer in specific tissues.

Myocardial protective effect of trilinolein: an antioxidant isolated from the medicinal plant Panax pseudoginseng.

In a previous study we demonstrated that trilinolein, a natural plant triacylglycerol, is a novel myocardial protective agent in vivo. The mechanism probably involves an antioxidant effect. This work investigated the mechanism of myocardial protection of trilinolein to determine if inhibition of calcium influx and alteration of activity of superoxide dismutase are involved. In isolated cardiomyocytes, pretreatment with trilinolein at a low concentration of 10(-9) M effectively reduced 45Ca2+ influx stimulated by hypoxia/normoxia by 34%. In isolated perfused rat heart subjected to 60 min global hypoxemia without reperfusion, pretreatment with 10(-7) M trilinolein for 15 min reduced infarct size by 37%. Assay of superoxide dismutase-mRNA by Northern blot analysis in in vivo rat heart subjected to 30 min ischaemia and 10 min reperfusion showed pretreatment with 10(-7) M trilinolein had a synergistic action with antioxidant systems preventing the rise in superoxide dismutase-mRNA. These results reconfirm the myocardial protection of trilinolein and suggest it may be related to antioxidant activity and inhibition of 45Ca2+ influx.

Complementation of hypopigmentation in p-mutant (pink-eyed dilution) mouse melanocytes by normal human P cDNA, and defective complementation by OCA2 mutant sequences.

Mutations in the P gene of humans and the homologous p-locus of mice, respectively, result in the homologous disorders oculocutaneous albinism type 2 (OCA2) and pink-eyed dilution. Although clearly required for melanin biosynthesis, the specific function of the P gene product, a melanosomal transmembrane protein expressed in melanocytes of the skin, hair, and eyes, is not yet known. Here we describe lines of immortal melanocytes and melanoblasts from mice of the null genotype p(cp)/p(25H). These p-null melanocytes were severely hypopigmented, although they and the melanoblasts expressed mRNAs for a number of melanosomal proteins. Proliferation of the p-null melanoblasts was normal. Both diploid and immortal p-null melanocytes grew more slowly than wild-type melanocytes, however, and were unusually susceptible to the antibiotic G418; these abnormalities were corrected by culture in high concentrations of L-tyrosine. Transfection of the p-null melanocytes with full-length normal human P cDNA resulted in complementation of deficient melanin biosynthesis and hypopigmentation. In contrast, transfection with mutant human P cDNAs containing amino acid substitutions (A481T, V443I) found in patients with OCA2 resulted in minimal or partial correction, consistent with the corresponding pigmentation phenotypes in patients with these mutations. These results demonstrate the utility of this model system for distinguishing true OCA2 mutations from nonpathologic polymorphisms and for quantitating the effect of these mutations on P function.

Characterization of the human full-length PTK7 cDNA encoding a receptor protein tyrosine kinase-like molecule closely related to chick KLG.

A 220-bp fragment of PTK7 cDNA was previously cloned from normal human melanocyte RNAs by means of the reverse transcription-polymerase chain reaction [Lee, S.-T., Strunk, K.M., and Spritz, R.A. (1993) Oncogene 8, 3403-3410]. We now report the cloning of the human full-length PTK7 cDNA and its characterization. The 1,070 amino acid PTK7 polypeptide deduced from the cDNA sequence constitutes receptor protein tyrosine kinase (RPTK), but has several unusual residues in some of the highly conserved tyrosine kinase motifs. PTK7 mRNA was expressed at the highest level in a human erythroleukemia cell line among tested samples, and at relatively high levels in liver, lung, pancreas, kidney, placenta, and melanocytes. Human PTK7 is 72% identical to chick KLG, suggesting that PTK7 is homologous or possibly orthologous to chick KLG, and that these represent a new subfamily of RPTKs.

Influence of a supplementary carbon source on biodegradation of pyridine by freely suspended and immobilized Pimelobacter sp..

The effect of the presence of supplementary glucose or acetate on the growth and pyridine-degrading activity of freely suspended and calcium-alginate-immobilized Pimelobacter sp. was investigated. Although the supplementary carbon sources could be degraded simultaneously with pyridine, Pimelobacter sp. exhibited a preference for pyridine over supplementary carbon sources. Thus, the pyridine-degrading activity of the freely suspended cells was not decreased significantly by the addition of either glucose (1.5-6 mM) or acetate (6-24 mM) to the pyridine (6-24 mM). In the semi-continuous immobilized cell culture, immobilized cells also exhibited a preference for pyridine over supplementary carbon sources and did not switch their substrate preference throughout the culture. Owing to a high cell concentration, the volumetric pyridine degradation rate at 24 mM pyridine in the immobilized cell culture was approximately six times higher than that in the freely suspended cell culture. Furthermore, the immobilized cells could be reused 16 times without losing their pyridine-degrading activity during the culture period tested. Taken together, the use of immobilized Pimelobacter sp. for the degradation of pyridine is quite feasible because of the preference for pyridine over supplementary carbon sources, the high volumetric pyridine degradation rate, and the reusability of immobilized cells.