Adherence and medication utilisation patterns of fixed-dose and free combination of angiotensin receptor blocker/thiazide diuretics among newly diagnosed hypertensive patients: a population-based cohort study.

OBJECTIVE: The study aimed to compare the adherence and persistence among newly diagnosed hypertensive patients using fixed-dose (FDC) and free combinations (FC) of angiotensin receptor blocker (ARB)/thiazide diuretic using Taiwan's National Health Insurance Research Database. METHODS: General linear regression and Kaplan-Meier analyses were used to estimate the impact of FDC on adherence [measured by medication possession ratio (MPR)] and persistence (time from day of initiation to treatment discontinuation) of ARB/thiazide diuretic. RESULTS: The adjusted MPRs were all significantly higher among FDC group compared with FC group (6 months: 66.55% vs. 63.86%; 1 year: 52.58% vs. 46.73%, 1.5 year: 46.30% vs. 38.07%; 2 year: 42.06% vs. 32.45%, all p < 0.001). Patients received FDC therapy were less likely to discontinue their therapy [adjusted hazard ratio (HR) 0.79, 95% CI = 0.74-0.85]. CONCLUSIONS: Our findings suggest that use of FDC is associated with higher adherence and persistence rates than use of FC in newly diagnosed hypertensive patients.

Slow development of ALS-like spinal cord pathology in mutant valosin-containing protein gene knock-in mice.

Pathological features of amyotrophic lateral sclerosis (ALS) include, in addition to selective motor neuron (MN) degeneration, the occurrence of protein aggregates, mitochondrial dysfunction and astrogliosis. SOD1 mutations cause rare familial forms of ALS and have provided the most widely studied animal models. Relatively recent studies implicating another protein, TDP-43, in familial and sporadic forms of ALS have led to the development of new animal models. More recently, mutations in the valosin-containing protein (VCP) gene linked to the human genetic disease, Inclusion Body Myopathy associated with Paget's disease of bone and frontotemporal dementia (IBMPFD), were found also to be associated with ALS in some patients. A heterozygous knock-in VCP mouse model of IBMPFD (VCP(R155H/+)) exhibited muscle, bone and brain pathology characteristic of the human disease. We have undertaken studies of spinal cord pathology in VCP(R155H/+) mice and find age-dependent degeneration of ventral horn MNs, TDP-43-positive cytosolic inclusions, mitochondrial aggregation and progressive astrogliosis. Aged animals (~24-27 months) show electromyography evidence of denervation consistent with the observed MN loss. Although these animals do not develop rapidly progressive fatal ALS-like disease during their lifespans, they recapitulate key pathological features of both human disease and other animal models of ALS, and may provide a valuable new model for studying events preceding onset of catastrophic disease.

Oral administration of an edible-mushroom-derived protein inhibits the development of food-allergic reactions in mice.

BACKGROUND: Food allergy is a common disease without effective treatment. Since strict elimination of food allergens may be difficult, strategies for effective intervention are urgently needed. OBJECTIVE: The aim was to investigate the prophylactic use of orally administrated FIP-fve, an immunomodulatory protein isolated from the edible mushroom Flammulina velutipes, in a murine model of food allergy. METHODS: BALB/c mice were immunized twice intraperitoneally with ovalbumin (OVA), at an interval of 2 weeks. Before and during each period of immunization, FIP-fve (200 microg per mouse) or phosphate-buffered saline was given orally every other day with a total of five doses. Then OVA-specific antibodies and cytokine profiles were determined. Subsequently, the mice were orally challenged with OVA. Symptoms of anaphylaxis, levels of plasma histamine, and histology of intestines were examined. RESULTS: Mice receiving oral FIP-fve treatment during sensitization to OVA had an impaired OVA-specific IgE response with a Th1-predominant cytokine profile. These mice were protected from systemic anaphylaxis-like symptoms induced by subsequent oral challenge with OVA. CONCLUSION: Oral administration of FIP-fve has a Th1-skewing effect on the development of the allergen-specific immune response, and may serve the purpose of immunoprophylaxis for food allergy and other allergic diseases.

Primary structure and function analysis of the Abrus precatorius agglutinin A chain by site-directed mutagenesis. Pro(199) Of amphiphilic alpha-helix H impairs protein synthesis inhibitory activity.

Abrus agglutinin (AAG), a low-toxicity protein from the plant Abrus precatorius, is less lethal than abrina (ABRa) in mice (LD(50) = 5 mg/kg versus 20 microg/kg of body weight). Nucleotide sequence analysis of a cDNA clone encoding full-length AAG showed an open reading frame with 1641 base pairs, corresponding to a 547-amino acid residue preproprotein containing a signal peptide and a linker region (two amino acid residues) between the AAG-A and AAG-B subunits. AAG had high homology to ABRa (77.8%). The 13 amino acid residues involved in catalytic function, which are highly conserved among abrins and ricins, were also conserved within AAG-A. The protein synthesis inhibitory activity of AAG-A (IC(50) = 3.5 nM) was weaker than that of ABRa-A (0.05 nM). Molecular modeling followed by site-directed mutagenesis showed that Pro(199) of AAG-A, located in amphiphilic helix H and corresponding to Asn(200) of ABRa-A, can induce bending of helix H. This bending would presumably affect the binding of AAG-A to its target sequence, GpApGpAp, in the tetraloop structure of the 28 S rRNA subunit and could be one of the major factors contributing to the relatively weak protein synthesis inhibitory activity and toxicity of AAG.

Dimerization of the N-terminal amphipathic alpha-helix domain of the fungal immunomodulatory protein from Ganoderma tsugae (Fip-gts) defined by a yeast two-hybrid system and site-directed mutagenesis.

A fungal immunomodulatory protein (Fip-gts) was purified from Ganoderma tsugae. The DNA encoding Fip-gts was isolated from a cDNA library of G. tsugae by reverse transcriptase-polymerase chain reaction. The complete amino acid sequence of Fip-gts, deduced from the nucleotide sequence of the cDNA, was the same as LZ-8 isolated from Ganodermn lucidum. Recombinant Fip-gts was expressed as a glutathione S-transferase fusion protein in Escherichia coli with a yield of 20 mg/liter of culture. Recombinant Fip-gts, purified to homogeneity, had the same blast formation stimulatory activity to human peripheral blood lymphocytes as native Fip-gts. The yeast two-hybrid system and site-directed mutagenesis were used to determine whether dimerization of Fip-gts occurred. Deletion analysis of the N-terminal amphipathic alpha-helix domain of Fip-gts identified a sequence of about 10 amino acids responsible for inducing immunomodulatory activity. Non-functional Fip-gts deletion mutants did not form dimers, whereas wild type Fip-gts did as determined by gel filtration. A mutant with deletions at Leu-5, Phe-7, and Leu-9 lost the amphipathic characteristics of the N-terminal domain and the ability to form dimers as well as its immunomodulatory activity. Fusion of Fip-gts with the DNA binding and the transactivation domains of GAL4 resulted in the activation of the lacZ activator gene, indicating the interaction of Fip-gts with it itself. The dimerization domain was further defined by analyzing the ability of the N-terminal 13 amino acids or Leu-5, Phe-7, and Leu-9 deletion mutants of Fip-gts to interact with the wild type Fip-gts. These experiments confirmed the N-terminal amphipathic alpha-helix as the dimerization domain and suggest that the dimerization of Fip-gts may play an important role in Fip-gts immunomodulatory activity.

Molecular cloning and expression of a fungal immunomodulatory protein, FIP-fve, from Flammulina velutipes.

FIP-fve, a fungal immunomodulatory protein, was isolated from the fruiting bodies of the edible mushroom, Flammulina velutipes. FIP-fve was shown to stimulate blast-forming activity of human peripheral blood lymphocytes and gene expression of interleukin-2, interferon-gamma and tumor necrosis factor-alpha. Repeated administration of FIP-fve to mice inhibits the Arthur and systemic anaphylaxis reactions. FIP-fve cDNA was cloned and sequenced, and the amino acid sequence of FIP-fve deduced from the nucleotide sequence is identical to that previously determined by protein sequencing. FIP-fve cDNA was amplified by polymerase chain reaction, ligated into the expression vector, pGEX-2T, and expressed in Escherichia coli as a fusion protein of glutathione S-transferase (GST) and FIP-fve. The GST-FIP-fve fusion protein was soluble, and the yield of recombinant FIP-fve was about 5 mg/L of induced culture. The recombinant FIP-fve was obtained by cleaving the GST-FIP-fve fusion protein with thrombin and purifing to homogeneity. The recombinant FIP-fve had about 50% of the immunomodulatory activity of the native FIP-fve.

Fip-vvo, a new fungal immunomodulatory protein isolated from Volvariella volvacea.

A new fungal immunomodulatory protein (Fip) has been purified from the edible mushroom, Volvariella volvacea, and designated Fip-vvo. Analysis of the purified protein by SDS/PAGE followed by Coomassie Blue staining demonstrated that Fip-vvo is a single polypeptide with an apparent molecular mass of 15 kDa. Periodic acid/Schiff staining showed that this single polypeptide lacks carbohydrates. Using an in vitro bioassay measuring blast-formation stimulatory activity, Fip-vvo was shown to stimulate the maximum proliferation of human peripheral blood lymphocytes at a concentration of 5 microg/ml. Fip-vvo was capable of agglutinating rat red blood cells. Neither haemagglutination nor mitogenic activities were inhibited by mono- or dimeric sugars. In vivo, repeat administration of Fip-vvo greatly reduced the production of BSA-induced Arthus reaction in mice, whereas little effect was observed on the prevention of systemic anaphylaxis reactions. The selectively enhanced transcriptional expression of interleukin (IL)-2, IL-4, interferon-gamma, tumour necrosis factor-alpha, lymphotoxin and IL-2 receptor by Fip-vvo was also demonstrated by reverse transcriptase-PCR. This finding suggests that Fip-vvo exerts its immunomodulatory effects via cytokine regulation. In addition, the complete amino acid sequence of Fip-vvo was obtained by direct protein sequencing. This protein consists of 112 amino acid residues with a blocked N-terminal end and has a calculated molecular mass of 12667 Da not including the N-terminal blocking group. By gel filtration analysis, Fip-vvo exhibited a molecular mass of 26 kDa for the native molecules in PBS. This result indicates that native Fip-vvo is most likely a non-covalently associated homodimeric molecule.

Two outbreaks of acute Tung Nut (Aleurites fordii) poisoning.

OBJECTIVE: Aleurites fordii, widely distributed in the Southeastern US, Taiwan, mainland China, Japan and India, is commonly known as Tung Nut, Tung Oil Tree or Chinawood Oil Tree. The seeds are the most toxic part. CASE REPORTS: We report two outbreaks of Aleurites fordii poisoning, occurring on November 27, 1992 and November 29, 1994. Thirty-five elementary school students and 29 senior high school students misidentified Aleurites fordii seeds as chestnuts and ingested variable amounts. METHODS: We conducted a survey by questionnaire to supplement the hospital record information. Simple descriptive statistics and Chi-square (Fisher's exact) tests were calculated. RESULTS: The three most common symptoms of the patients were vomiting, abdominal pain and diarrhea. The more serious clinical presentations occurred in younger victims. Our information suggests that food attenuates intestinal irritation perhaps by delaying absorption of the toxic principle. With symptomatic treatment, all of the symptoms and signs subsided within one to two days. CONCLUSIONS: Aleurites fordii can be cultivated and is easily accessible to the community and schools. Public education about the toxicity of Tung Nut seeds in areas of ready availability may reduce the chance of misidentification and subsequent poisoning.

A new fungal immunomodulatory protein, FIP-fve isolated from the edible mushroom, Flammulina velutipes and its complete amino acid sequence.

A new fungal immunomodulatory protein (FIP-fve) has been isolated and purified from the edible golden needle mushroom (Flammulina velutipes). The apparent molecular mass of FIP-fve determined by SDS/PAGE agrees well with the value of 12704 Da calculated from its amino acid composition and sequence. The complete amino acid sequence of FIP-fve was elucidated by protein sequencing techniques. FIP-fve consists of 114 amino acid residues with an acetylated amino end, and lacks methionine, half-cystine and histidine residues. FIP-fve was able to hemagglutinate human red blood cells. The immunomodulatory activity of FIP-fve was demonstrated by its stimulatory activity toward human peripheral blood lymphocytes, and its suppression of systemic anaphylaxis reactions and local swelling of mouse footpads. FIP-fve was found to enhance the transcriptional expression of interleukin-2 and interferon-gamma.