Identification and management of contraindicated drug-drug interactions through pharmaceutical care programs: Experience in direct-acting antivirals therapy.

BACKGROUND/PURPOSE: To investigate the impact of pharmaceutical care programs for the management of contraindicated drug-drug interactions (DDIs) in direct-acting antivirals (DAAs) therapy. METHODS: A prospective observational study was performed at Dalin Tzu Chi Hospital between January 2018 and December 2019. Pharmacists screened DDIs for all hepatitis C patients before DAA therapy. The study outcome included the frequency of contraindicated DDIs, acceptance rate, and cost avoidance of the pharmaceutical care program. RESULTS: A total of 1053 patients were enrolled in the study, with a mean age of 67.1 ± 11.9 years. Most patients received therapy with sofosbuvir/ledipasvir (37.1%; n = 391), elbasvir/grazoprevir (23.8%; n = 251), or glecaprevir/pibrentasvir (21.1%; n = 222). A total of 796 (75.6%) patients received at least one co-medication, with the average number of co-medications being 5.2 per patient (SD: 4.4/patient). In total, 1356 DDIs were identified, with the average DDIs per patient of 1.3 (SD: 1.7). For patients with contraindicated DDIs (2%, n = 102), statins and amiodarone were the most common co-medications. Physicians often accepted pharmacists' recommendations (acceptance rate of 72.5%) or withheld co-medication to avoid severe adverse drug events (ADEs). The estimated cost avoidance of preventable ADEs was USD 14,033 for contraindicated DDIs with pharmaceutical care programs. CONCLUSION: The implementation of pharmaceutical care programs in DAA therapy provides a favorable outcome and substantial cost avoidance.

Perturbation of cytosolic calcium by 2-aminoethoxydiphenyl borate and caffeine affects zebrafish myofibril alignment.

The objective of the current study was to investigate the effects of Ca(2+) levels on myofibril alignment during zebrafish embryogenesis. To investigate how altered cytoplasmic Ca(2+) levels affect myofibril alignment, we exposed zebrafish embryos to 2-aminothoxyldiphenyl borate (2-APB; an inositol 1,4,5-trisphosphate receptor inhibitor that reduces cytosolic Ca(2+) levels) and caffeine (a ryanodine receptor activator that enhances cytosolic Ca(2+) levels). The results demonstrated that the most evident changes in zebrafish embryos treated with 2-APB were shorter body length, curved trunk and malformed somite boundary. In contrast, such malformed phenotypes were evident neither in untreated controls nor in caffeine-treated embryos. Subtle morphological changes, including changes in muscle fibers, F-actin and ultrastructures were easily observed by staining with specific monoclonal antibodies (F59 and α-laminin), fluorescent probes (phalloidin) and by transmission electron microscopy. Our data suggested that: (1) the exposure to 2-APB and/or caffeine led to myofibril misalignment; (2) 2-APB-treated embryos displayed split and short myofibril phenotypes, whereas muscle fibers from caffeine-treated embryos were twisted and wavy; and (3) zebrafish embryos co-exposed to 2-APB and caffeine resulted in normal myofibril alignment. In conclusion, we proposed that cytosolic Ca(2+) is important for myogenesis, particularly for myofibril alignment.

Plant chitinase III Ziz m 1 stimulates multiple cytokines, most predominantly interleukin-13, from peripheral blood mononuclear cells of latex-fruit allergic patients.

BACKGROUND: Indian jujube is a fruit abundantly cultivated in Taiwan. Its major allergen in latex-fruit syndrome is Ziz m 1 of the chitinase III family. The Ziz m 1 Pichia (rZiz m 1-P) has chitinase activity but not Ziz m 1 E. coli (rZiz m 1-E). OBJECTIVE: This study examined whether plant chitinase III, using rZiz m 1-P and rZiz m 1-E, can stimulate allergic inflammation similar to that of mammalian chitinases. METHODS: Five patients allergic to latex-Indian jujube and five nonallergic controls were evaluated. Their peripheral blood mononuclear cells (PBMC) were cultured with rZiz m 1-E or rZiz m 1-P and pulsed with phorbol 12-myristate 13-acetate. Eleven cytokines were measured by FlowCytomix human Th1/Th2 plex kit and interleukin (IL)-13 by sandwich enzyme-linked immunosorbent assay (ELISA). RESULTS: Interleukin-13 significantly increased in rZiz m 1-P stimulated PBMC of allergic subjects but was undetectable when stimulated with rZiz m 1-E. The stimulation index significantly increased in IL-13 (380.6 ± 77.33 vs 13.70 ± 6.92), IL-5 (6.70 ± 0.59 vs 0.70 ± 0.37), IL-1ß (32.70 ± 0.83 vs 2.10 ± 1.29), and tumor necrosis factor beta (TNF-ß) (17.10 ± 2.66 vs 1.50 ± 0.66) between allergic and nonallergic subjects after rZiz m 1-P stimulation. There was no difference in terms of IL-2, IFN-γ, IL-8, and TNF-α production. CONCLUSIONS: The biological function of chitinase activity is required for Ziz m 1 to induce a Th2-specific immune response. This is the first report on PBMC responses of latex-fruit syndrome subjects toward an active exogenous plant class III chitinase that can stimulate multiple cytokines, especially IL-13, from allergic subjects. This implies the role of cross-reactive food allergens in propagating allergic inflammation among allergic subjects.

Demonstration of expression of a neuropeptide-encoding gene in crustacean hemocytes.

Crustacean hyperglycemic hormone (CHH) was originally identified in a neuroendocrine system-the X-organ/sinus gland complex. In this study, a cDNA (Prc-CHH) encoding CHH precursor was cloned from the hemocyte of the crayfish Procambarus clarkii. Analysis of tissues by a CHH-specific enzyme-linked immunosorbent assay (ELISA) confirmed the presence of CHH in hemocytes, the levels of which were much lower than those in the sinus gland, but 2 to 10 times higher than those in the thoracic and cerebral ganglia. Total hemocytes were separated by density gradient centrifugation into layers of hyaline cell (HC), semi-granular cell (SGC), and granular cell (GC). Analysis of extracts of each layer using ELISA revealed that CHH is present in GCs (202.8±86.7 fmol/mg protein) and SGCs (497.8±49.4 fmol/mg protein), but not in HCs. Finally, CHH stimulated the membrane-bound guanylyl cyclase (GC) activity of hemocytes in a dose-dependent manner. These data for the first time confirm that a crustacean neuropeptide-encoding gene is expressed in cells essential for immunity and its expression in hemocytes is cell type-specific. Effect of CHH on the membrane-bound GC activity of hemocyte suggests that hemocyte is a target site of CHH. Possible functions of the hemocyte-derived CHH are discussed.

Poly(allylguanidine)-Coated Surfaces Regulate TGF-ß in Glioblastoma Cells to Induce Apoptosis via NF-κB Pathway Activation.

Polycationic biomaterials are currently widely applied in neuronal cell cultures to promote cell adhesion and viability. However, polycations generally have cytotoxic properties that limit their application in the field of biomaterials. In this study, we examined the use of a novel polycation poly(allylguanidine) (PAG), which contains a guanidine group in the side chain and a structure similar to poly(allylamine hydrochloride) (PAH), an example of another commonly used polycation. Our findings showed that exposure to PAG induced apoptosis in glioblastoma (GBM) cells, while exposure to PAH induced necrosis. Compared to control groups, the PAG coating significantly limited the proliferation of GBM8901 in vitro and in vivo. Furthermore, GBM8901 cells exposed to the PAG coating exhibited increased levels of phospho-p65 and phosphor-IκB, implying that GBM8901 cells underwent apoptotic cell death via the NF-κB pathway by the regulation of TGF-ß. This result was further confirmed to be consistent with the experimental results from western blot protein analysis and apoptosis/necrosis assays. These findings indicate that the polycation PAG has the potential to not only suppress the proliferation of GBM8901 cancer cells but also improve the neural viability and promote the differentiation of neural stem/precursor cells into mature neurons. In conclusion, biomaterials such as PAG act as extremely potent options for applications in the treatment of pathological conditions such as brain cancer.

Specific IgE and IgG responses and cytokine profile in subjects with allergic reactions to biting midge Forcipomyia taiwana.

BACKGROUND: Forcipomyia taiwana is a tiny blood-sucking midge whose habitat covers large parts of Taiwan and southern China. Female midges bite during the day, causing intense pruritus and swelling in allergic individuals. In this study, we investigated the immune responses of different allergic reactions to midge bites. METHODS: F. taiwana (midge)-specific IgE, -IgG and -IgG subclasses were examined by ELISA in 62 human subjects. Peripheral blood mononuclear cells (PBMC) from 6 subjects with solely delayed reactions (SDR) to midge bites and 6 nonallergic controls (NAC) were cultured with midge extract at various time points and assayed. Proliferation of PBMC was measured by MTT assay. Expression of cytokine mRNA was measured by real-time PCR and protein levels by cytometric bead immunoassay or ELISA. Protease activity in midge extract was determined by the Azocoll method. RESULTS: Midge-specific IgE among subjects with an immediate reaction were significantly elevated compared to SDR and NAC subjects. There were no differences in the level of midge-specific-IgG, -IgG(1), -IgG(2), -IgG(3) and -IgG(4) among subjects with different biting reactions. Midge extract elicited significantly more PBMC proliferation, higher expression of IFN-gamma, IL-10, IL-6 and TNF-alpha in SDR subjects than in NAC. Protease activity was detected in midge extract. Protease inhibitors E64 and pepstatin suppressed midge-extract-induced IL-8 production. CONCLUSIONS: Our results suggest that an immediate reaction to midge bites is IgE-mediated. IFN-gamma, IL-6 and TNF-alpha are involved in delayed reactions to midge bites. A protease-activated pathway may also be involved in the intense, itchy reactions to midge bites.

Transgenic zebrafish line with over-expression of Hedgehog on the skin: a useful tool to screen Hedgehog-inhibiting compounds.

We generated a transgenic line Tg(k18:shh:RFP) with overexpression of Sonic hedgehog in the skin epidermis. By 5 day-post-fertilization (dpf), many epidermal lesions were clearly observed, including a swollen yolk sac, epidermis growth malformation around the eyes and at the basement of the pectoral fins. Skin histology revealed embryos derived from Tg(k18:shh:RFP) displayed an elevated Nuclear/Cytoplasmic ratio and pleomorphic nuclei compared to their wild type littermates, suggesting the abnormal growth pattern on the epidermis of Tg(k18:shh:RFP) embryos were dysplasia. Later (by 7 dpf), Tg(k18:shh:RFP) embryos displayed broader pectoral fins which are similar to the polydactyly phenotypes of Nevoid basal cell carcinoma syndrome (NBCCS)/Gorlin patients and polydactylous mice. In addition, treatment with cyclopamine is able to enhance and prolong the survival rates and survival durations of Tg(k18:shh:RFP) embryos. In conclusion, this unique Tg(k18:shh:RFP) fish line, should be an excellent experimental animal for screening for a lower toxicity level of the new Hh-inhibitor and can even be used as a new anti-cancer drug-screening platform.

Preparation of aligned poly(glycerol sebacate) fibrous membranes for anisotropic tissue engineering.

The use of fibrous scaffolds for tissue repair or regeneration is advantageous for its microstructure similar to that of the native ECM. Aligned fibrous scaffold, in particular, can be used to manipulate cell alignment and hence the microstructure of the resultant tissue. In our previous study, nanofibers consisting of solely poly(glycerol sebacate) (PGS) have been successfully fabricated using core-shell coaxial electrospinning followed by curing and subsequent shell removal. When we tried to fabricate aligned PGS fibrous membranes by collecting the electrospun fibers on a rapidly rotating drum, however, loss of fibrous structure was observed upon curing. This might be due to the broken fibers that were collected under tension; the core PGS prepolymer that melts at high temperature could leak from the broken ends during curing. In this study, attempts were made to reduce the possibility of the fiber breakage. At each stage of preparation, fiber morphology was examined by SEM and fiber compositions were verified by Fourier transform infrared spectroscopy and differential scanning calorimetry. Mechanical properties of the aligned PGS fibrous membrane were evaluated by uniaxial tensile testing both in parallel and perpendicular to the principal fiber direction. SEM images showed that fibrous morphology was better preserved upon the adjustment of the shell composition and the rotational speed of the collector drum. The final PGS fibers remained to be aligned although the alignment was less strong than that of as-spun core-shell fibers. The aligned PGS fibrous membrane exhibited anisotropic mechanical properties with Young's modulus in parallel and perpendicular to the principal fiber direction being 0.98 ±â€¯0.04 MPa and 0.52 ±â€¯0.02 MPa, respectively. The aligned PGS fibrous membrane was capable of guiding the orientation of cultured cells and therefore has the potential to be used to fabricate structurally anisotropic tissue-engineered constructs.

Molecular cloning and differential expression pattern of two structural variants of the crustacean hyperglycemic hormone family from the mud crab Scylla olivacea.

Two full-length cDNA sequences encoding a crustacean hyperglycemic hormone (CHH) precursor were cloned from tissues of the mud crab Scylla olivacea. Sco-CHH (S. olivacea CHH) was cloned from eyestalk ganglia, whereas Sco-CHH-L (S. olivacea CHH-like peptide) was cloned from extra-eyestalk tissues (pericardial organ and thoracic ganglia). Each conceptually translated precursor is expected to be processed into a signal peptide, a CHH precursor-related peptide (CPRP), and a mature CHH or CHH-like peptide. The two precursors are identical in amino acid sequence through the 40th residue of the mature peptide, but different from each other substantially in the C-terminus. Both CHH variants contain the six highly conserved cysteine residues characteristic of the CHH family peptides, and share higher sequence identities with other brachyuran CHH sequences than with those of other taxonomic groups. As determined by reverse transcription-polymerase chain reaction (RT-PCR), the transcripts of Sco-CHH and Sco-CHH-L were present in eyestalk ganglia and several extra-eyestalk tissues (the thoracic ganglia, pericardial organ, brain, circumesophageal connectives, and gut). Sco-CHH was the predominant form in eyestalk ganglia, while Sco-CHH-L was the predominant form in several extra-eyestalk tissues. Neither transcript was expressed in the muscle, hepatopancreas, ovary, testis, heart, or gill. Antisera were raised against synthetic peptides corresponding to a stretch of sequence-specific to the C-terminus of Sco-CHH or Sco-CHH-L. Western blot analyses of tissues expressing Sco-CHH and Sco-CHH-L detected a Sco-CHH immunoreactive protein in the sinus gland, and a Sco-CHH-L immunoreactive protein in the pericardial organ. Immunohistochemical analyses of the eyestalk ganglia localized both Sco-CHH and Sco-CHH-L immunoreactivity to the sinus gland, and only Sco-CHH immunoreactivity to the X-organ somata; analyses of the pericardial organs also localized both Sco-CHH and Sco-CHH-L immunoreactivity to the anterior and posterior bars, as well as to longitudinal trunks joining the two bars. The combined data provided supporting evidence that Sco-CHH and Sco-CHH-L are co-localized in the same tissue.

Development of a whole-organism model to screen new compounds for sun protection.

We used zebrafish as a whole-organism model to screen new compounds for sun protection activity. First of all, we designed a series of UVB exposure experiments and recorded the phenotypic changes of zebrafish embryos. Results showed that 100 mJ/cm(2) of UVB given six times separated by 30 min intervals is the best condition. Fin malformation (reduced and/or absent fin) phenotypes are the most evident consequences after exposure to UVB. Each fin was affected by UVB, including pelvic, ventral, caudal, and dorsal fin, but pelvic fin seemed to be the most sensitive target after UVB exposure. We furthermore carried out "prevention" and "treatment" experiments using green tea extract and/or (-)-epigallocatechin (EGCG) to test this whole-organism model by observing the morphological changes of all fins (especially pelvic fin) after UVB exposure. Effects of UVB, green tea extract and EGCG on fin development were assessed using the Kaplan-Meier analysis, log-rank test and Cox proportional hazards regression. Results showed that a zebrafish pelvic fin in the UVB + green tea (treatment) group is 5.51 (range from 2.39 to 14.90) times, one in the UVB + green tea (prevention) group is 7.04 (range from 3.11 to 18.92) times, and one in the 25 ppm of EGCG (prevention) group is 22.19 (range from 9.40 to 61.50) times more likely to return to normal fin than one in the UVB only group. On the basis of these observations, we believe this model is effective for screening the higher stability and lower toxicity of new compounds, such as small chemicals which are derivative from EGCG or other dietary agents for sun protection.

A hospital-based screening study of latex allergy and latex sensitization among medical workers in Taiwan.

BACKGROUND AND PURPOSE: Natural rubber latex is the most important occupational allergen among medical workers, and remains a significant occupational health issue in Taiwan. We conducted this large-scale hospital-based screening study to understand the incidence of latex allergy and latex sensitization among medical workers in Taiwan over the past 5 years. METHODS: 1253 medical workers were enrolled in this study. Subjects were interviewed using a screening questionnaire. Skin prick testing with commercial latex extract was performed for 1139 of the subjects. RESULTS: 152 subjects (12%) had positive latex skin prick test, suggesting that they had been sensitized to latex proteins. Seventy nine subjects (6%) had immediate allergic reactions to latex products. The prevalence of contact hand dermatitis from latex gloves was 35%. The intensive care unit and medical laboratory department accounted for the highest prevalence of allergy among all hospital departments. Most subjects developed immediate latex allergy by 9000 h of total latex exposure. The prevalence of positive latex skin prick test increased with increasing duration of latex exposure. CONCLUSIONS: Latex allergy continues to be an important occupational allergy among medical workers in Taiwan. The addition of a routine screening examination in medical employees' health check-ups will help in the early identification of sensitized cases and facilitate preventive strategies.