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Metabolic glycan labeling and chemoselective functionalization of native biomaterials

Ren X, Evangelista-Leite D, Wu T, Rajab TK, Moser PT, Kitano K, et al.

Biomaterials, 2018 11;182:127-134.
PMID: 30118980 DOI: 10.1016/j.biomaterials.2018.08.012

Decellularized native extracellular matrix (ECM) biomaterials are widely used in tissue engineering and have reached clinical application as biomesh implants. To enhance their regenerative properties and postimplantation performance, ECM biomaterials could be functionalized via immobilization of bioactive molecules. To facilitate ECM functionalization, we developed a metabolic glycan labeling approach using physiologic pathways to covalently incorporate click-reactive azide ligands into the native ECM of a wide variety of rodent tissues and organs in vivo, and into the ECM of isolated rodent and porcine lungs cultured ex vivo. The incorporated azides within the ECM were preserved after decellularization and served as chemoselective ligands for subsequent bioconjugation via click chemistry. As proof of principle, we generated alkyne-modified heparin, immobilized it onto azide-incorporated acellular lungs, and demonstrated its bioactivity by Antithrombin III immobilization and Factor Xa inhibition. The herein reported metabolic glycan labeling approach represents a novel platform technology for manufacturing click-reactive native ECM biomaterials, thereby enabling efficient and chemoselective functionalization of these materials to facilitate tissue regeneration and repair.

Matched MeSH terms: Swine
Design, characterization, and evaluation of intranasal delivery of ropinirole-loaded mucoadhesive nanoparticles for brain targeting

Jafarieh O, Md S, Ali M, Baboota S, Sahni JK, Kumari B, et al.

Drug Dev Ind Pharm, 2015;41(10):1674-81.
PMID: 25496439 DOI: 10.3109/03639045.2014.991400

Parkinson disease (PD) is a common, progressive neurodegenerative disorder, characterized by marked depletion of striatal dopamine and degeneration of dopaminergic neurons in the substantia nigra.

Matched MeSH terms: Swine
Fulltext Prevalence and risk factors of Japanese encephalitis virus (JEV) in livestock and companion animal in high-risk areas in Malaysia

Kumar K, Arshad SS, Selvarajah GT, Abu J, Toung OP, Abba Y, et al.

Trop Anim Health Prod, 2018 Apr;50(4):741-752.
PMID: 29243139 DOI: 10.1007/s11250-017-1490-6

Japanese encephalitis (JE) is vector-borne zoonotic disease which causes encephalitis in humans and horses. Clinical signs for Japanese encephalitis virus (JEV) infection are not clearly evident in the majority of affected animals. In Malaysia, information on the prevalence of JEV infection has not been established. Thus, a cross-sectional study was conducted during two periods, December 2015 to January 2016 and March to August in 2016, to determine the prevalence and risk factors in JEV infections among animals and birds in Peninsular Malaysia. Serum samples were harvested from the 416 samples which were collected from the dogs, cats, water birds, village chicken, jungle fowls, long-tailed macaques, domestic pigs, and cattle in the states of Selangor, Perak, Perlis, Kelantan, and Pahang. The serum samples were screened for JEV antibodies by commercial IgG ELISA kits. A questionnaire was also distributed to obtain information on the animals, birds, and the environmental factors of sampling areas. The results showed that dogs had the highest seropositive rate of 80% (95% CI: ± 11.69) followed by pigs at 44.4% (95% CI: ± 1.715), cattle at 32.2% (95% CI: ± 1.058), birds at 28.9% (95% CI: ± 5.757), cats at 15.6% (95% CI: ± 7.38), and monkeys at 14.3% (95% CI: ± 1.882). The study also showed that JEV seropositivity was high in young animals and in areas where mosquito vectors and migrating birds were prevalent.

Matched MeSH terms: Swine
Fulltext Bartonella and Rickettsia in arthropods from the Lao PDR and from Borneo, Malaysia

Kernif T, Socolovschi C, Wells K, Lakim MB, Inthalad S, Slesak G, et al.

Comp Immunol Microbiol Infect Dis, 2012 Jan;35(1):51-7.
PMID: 22153360 DOI: 10.1016/j.cimid.2011.10.003

Rickettsioses and bartonelloses are arthropod-borne diseases of mammals with widespread geographical distributions. Yet their occurrence in specific regions, their association with different vectors and hosts and the infection rate of arthropod-vectors with these agents remain poorly studied in South-east Asia. We conducted entomological field surveys in the Lao PDR (Laos) and Borneo, Malaysia by surveying fleas, ticks, and lice from domestic dogs and collected additional samples from domestic cows and pigs in Laos. Rickettsia felis was detected by real-time PCR with similar overall flea infection rate in Laos (76.6%, 69/90) and Borneo (74.4%, 268/360). Both of the encountered flea vectors Ctenocephalides orientis and Ctenocephalides felis felis were infected with R. felis. The degrees of similarity of partial gltA and ompA genes with recognized species indicate the rickettsia detected in two Boophilus spp. ticks collected from a cow in Laos may be a new species. Isolation and further characterization will be necessary to specify it as a new species. Bartonella clarridgeiae was detected in 3/90 (3.3%) and 2/360 (0.6%) of examined fleas from Laos and Borneo, respectively. Two fleas collected in Laos and one flea collected in Borneo were co-infected with both R. felis and B. clarridgeiae. Further investigations are needed in order to isolate these agents and to determine their epidemiology and aetiological role in unknown fever in patients from these areas.

Matched MeSH terms: Swine
Quantitative autoradiographic studies of relaxin binding in rat atria, uterus and cerebral cortex: characterization and effects of oestrogen treatment

Tan YY, Wade JD, Tregear GW, Summers RJ

Br J Pharmacol, 1999 May;127(1):91-8.
PMID: 10369460

The binding characteristics of the relaxin receptor in rat atria, uterus and cortex were studied using a [33P]-labelled human gene 2 relaxin (B33) and quantitative receptor autoradiography. The binding kinetics of [33P]-human gene 2 relaxin (B33) were investigated in slide-mounted rat atrial sections. The binding achieved equilibrium after 60 min incubation at room temperature (23+/-1 degrees C) and dissociated slowly. The association and dissociation rate constants were 4.31+/-0.34x10(8) M(-1) x min(-1) and 1.55+/-0.38x10(-3) min(-1) respectively. Thus, the kinetic dissociation constant was 3.46+/-0.59 pM. Binding was saturable to a single population of non-interacting sites throughout atria, in uterine myometrium and the 5th layer of cerebral cortex. The binding affinities (pK(D)) of [33P]-human gene 2 relaxin (B33) were 8.92+/-0.09 in atrial myocardium and 8.79+/-0.04 in cerebral cortex of male rats, and 8.79+/-0.10 in uterine myometrium. Receptor densities in the cerebral cortex and atria were higher than in uterine myometrium, indicating that relaxin also has important roles in non-reproductive tissues. In male rats, treatment with 17beta-oestradiol (20 microg in 0.1 ml sesame oil s.c., 18-24 h) significantly decreased the density of relaxin receptors in atria and cerebral cortex. Identical treatment in female rats had no significant effect in atria and cerebral cortex, but it significantly increased the density of relaxin receptors in uterine myometrium. Relaxin binding was competitively displaced by porcine and rat native relaxins. Porcine native relaxin binds to the relaxin receptor in male rat atria (8.90+/-0.02), and cerebral cortex (8.90+/-0.03) and uterine myometrium (8.89+/-0.03) with affinities not significantly different from human gene 2 (B33) relaxin. Nevertheless, rat relaxin binds to the receptors with affinities (8.35+/-0.09 in atria, 8.22+/-0.07 in cerebral cortex and 8.48+/-0.06 in uterine myometrium) significantly less than human gene 2 (B33) and porcine relaxins. Quantitative receptor autoradiography is the method of choice for measurement of affinities and densities of relaxin receptor in atria, uterine myometrium and cerebral cortex. High densities were found in all these tissues. 17beta-oestradiol treatment produced complex effects where it increased the densities of relaxin receptors in uterus but decreased those in atria and cerebral cortex of the male rats, and had no effect on the atria and cerebral cortex of the female rats.

Matched MeSH terms: Swine
Fulltext Comparison of serum and oral fluid antibody responses after vaccination with a modified live (MLV) porcine reproductive and respiratory syndrome virus (PPRSV) vaccine in PRRS endemic farms

Kuiek AM, Ooi PT, Yong CK, Ng CF

Trop Anim Health Prod, 2015 Oct;47(7):1337-42.
PMID: 26070293 DOI: 10.1007/s11250-015-0868-6

Porcine reproductive and respiratory syndrome (PRRS) is a disease that is both highly contagious and of great economic importance in Malaysia. Therefore, reliable and improved diagnostic methods are needed to facilitate disease surveillance. This study compared PRRSV antibody responses in oral fluid versus serum samples following PRRS modified live (MLV) vaccination using commercial antibody ELISA kits (IDEXX Laboratories, Inc.). The study involved two pig farms located in Perak and Selangor, Malaysia. Both farms were vaccinated with PRRS MLV 1 month prior to sample collection. Thirty-five animals were used as subjects in each farm. These 35 animals were divided into 7 different categories: gilts, young sows, old sows, and four weaner groups. Oral fluid and serum samples were collected from these animals individually. In addition, pen oral fluid samples were collected from weaner groups. The oral fluid and serum samples were tested with IDEXX PRRS Oral Fluid Antibody Test Kit and IDEXX PRRS X3 Antibody Test Kit, respectively. The results were based on sample to positive ratio (S/P ratio of the samples). Results revealed a significant and positive correlation between serum and oral fluid samples for both farm A (p = 0.0001, r = 0.681) and farm B (p = 0.0001, r = 0.601). In general, oral fluids provided higher S/P results than serum, but the patterns of response were highly similar, especially for the sow groups. Thus, the use of oral fluids in endemic farms is effective and economical, particularly for large herds. In conclusion, the authors strongly recommend the use of oral fluids for PRRS monitoring in endemic farms.

Matched MeSH terms: Swine
Late clinical and magnetic resonance imaging follow up of Nipah virus infection

Lim CC, Lee WL, Leo YS, Lee KE, Chan KP, Ling AE, et al.

J Neurol Neurosurg Psychiatry, 2003 Jan;74(1):131-3.
PMID: 12486285

The Nipah virus is a newly identified paramyxovirus responsible for an outbreak of fatal encephalitis in Malaysia and Singapore. This paper reports the follow up clinical and magnetic resonance imaging findings in 22 affected subjects. Of 13 patients with encephalitis, one died, one was lost to follow up, and seven recovered. Among the four remaining patients, one had residual sixth nerve palsy, another suffered from severe clinical depression, and a third patient had evidence of retinal artery occlusion. One patient with delayed onset Horner syndrome had a single lesion in the cervical spinal cord. The brain magnetic resonance findings were stable or improved in nine patients over 18 months of follow up. Among a second group of nine asymptomatic seropositive abattoir workers, magnetic resonance examination in seven subjects revealed discrete small lesions in the brain; similar to those detected in encephalitis patients. These findings suggest that in addition to encephalitis, the newly discovered Nipah virus affects the spinal cord and the retina. Late clinical and radiological findings can occur in Nipah virus infections as with other paramyxoviruses.

Matched MeSH terms: Swine
Purification of the extra-cellular domain of Nipah virus glycoprotein produced in Escherichia coli and possible application in diagnosis

Eshaghi M, Tan WS, Chin WK, Yusoff K

J Biotechnol, 2005 Mar 30;116(3):221-6.
PMID: 15707682

The glycoprotein (G) of Nipah virus (NiV) is important for virus infectivity and induction of the protective immunity. In this study, the extra-cellular domain of NiV G protein was fused with hexahistidine residues at its N-terminal end and expressed in Escherichia coli. The expression under transcriptional regulation of T7 promoter yielded insoluble protein aggregates in the form of inclusion bodies. The inclusion bodies were solubilized with 8 M urea and the protein was purified to homogeneity under denaturing conditions using nickel-nitrilotriacetic acid (Ni-NTA) affinity chromatography. The denatured protein was renatured by gradual removal of the urea. Light scattering analysis of the purified protein showed primarily monodispersity. The purified protein showed significant reactivity with the antibodies present in the sera of NiV-infected swine, as demonstrated in Western blot analysis and enzyme-linked immunosorbent assay (ELISA). Taken together, the data indicate the potential usefulness of the purified G protein for structural or functional studies and the development of immunoassay for detection of the NiV antibodies.

Matched MeSH terms: Swine
Fulltext Indirect ELISA based on Hendra and Nipah virus proteins for the detection of henipavirus specific antibodies in pigs

Fischer K, Diederich S, Smith G, Reiche S, Pinho Dos Reis V, Stroh E, et al.

PLoS One, 2018;13(4):e0194385.
PMID: 29708971 DOI: 10.1371/journal.pone.0194385

Hendra virus (HeV) and Nipah virus (NiV) belong to the genus Henipavirus in the family Paramyxoviridae. Henipavirus infections were first reported in the 1990's causing severe and often fatal outbreaks in domestic animals and humans in Southeast Asia and Australia. NiV infections were observed in humans in Bangladesh, India and in the first outbreak in Malaysia, where pigs were also infected. HeV infections occurred in horses in the North-Eastern regions of Australia, with singular transmission events to humans. Bats of the genus Pteropus have been identified as the reservoir hosts for henipaviruses. Molecular and serological indications for the presence of henipa-like viruses in African fruit bats, pigs and humans have been published recently. In our study, truncated forms of HeV and NiV attachment (G) proteins as well as the full-length NiV nucleocapsid (N) protein were expressed using different expression systems. Based on these recombinant proteins, Enzyme-linked Immunosorbent Assays (ELISA) were developed for the detection of HeV or NiV specific antibodies in porcine serum samples. We used the NiV N ELISA for initial serum screening considering the general reactivity against henipaviruses. The G protein based ELISAs enabled the differentiation between HeV and NiV infections, since as expected, the sera displayed higher reactivity with the respective homologous antigens. In the future, these assays will present valuable tools for serosurveillance of swine and possibly other livestock or wildlife species in affected areas. Such studies will help assessing the potential risk for human and animal health worldwide by elucidating the distribution of henipaviruses.

Matched MeSH terms: Swine
Immune responses in mice and pigs after oral vaccination with rabies virus vectored Nipah disease vaccines

Shuai L, Ge J, Wen Z, Wang J, Wang X, Bu Z

Vet Microbiol, 2020 Feb;241:108549.
PMID: 31928698 DOI: 10.1016/j.vetmic.2019.108549

Nipah virus (NiV) is a re-emerging zoonotic pathogen that causes high mortality in humans and pigs. Oral immunization in free-roaming animals is one of the most practical approaches to prevent NiV pandemics. We previously generated a recombinant rabies viruses (RABV) Evelyn-Rokitnicki-Abelseth (ERA) strain, rERAG333E, which contains a mutation from arginine to glutamic acid at residue 333 of glycoprotein (G333E) and serves as an oral vaccine for dog rabies. In this study, we generated two recombinant RABVs, rERAG333E/NiVG and rERAG333E/NiVF, expressing the NiV Malaysian strain attachment glycoprotein (NiV-G) or fusion glycoprotein (NiV-F) gene based on the rERAG333E vector platform. Both rERAG333E/NiVG and rERAG333E/NiVF displayed growth properties similar to those of rERAG333E and caused marked syncytia formation after co-infection in BSR cell culture. Adult and suckling mice intracerebrally inoculated with the recombinant RABVs showed NiV-G and NiV-F expression did not increase the virulence of rERAG333E. Oral vaccination with rERAG333E/NiVG either singularly or combined with rERAG333E/NiVF induced significant NiV neutralizing antibody against NiV and RABV, and IgG to NiV-G or NiV-F in mice and pigs. rERAG333E/NiVG and rERAG333E/NiVF thus appeared to be suitable candidates for further oral vaccines for potential animal targets in endemic areas of NiV disease and rabies.

Matched MeSH terms: Swine
Fulltext Therapeutic Potential of Bama Pig Adipose-Derived Mesenchymal Stem Cells for the Treatment of Carbon Tetrachloride-Induced Liver Fibrosis

Wu X, Zhang S, Lai J, Lu H, Sun Y, Guan W

Exp Clin Transplant, 2020 12;18(7):823-831.
PMID: 33349209 DOI: 10.6002/ect.2020.0108

OBJECTIVES: Liver fibrosis is inevitable in the healing process of liver injury. Liver fibrosis will develop into liver cirrhosis unless the damaging factors are removed. This study investigated the potential therapy of Bama pig adipose-derived mesenchymal stem cells in a carbon tetrachloride-induced liver fibrosis Institute of Cancer Research strain mice model.
MATERIALS AND METHODS: Adipose-derived mesenchymal stem cells were injected intravenously into the tails of mice of the Institute of Cancer Research strain that had been treated with carbon tetrachloride for 4 weeks. Survival rate, migration, and proliferation of adipose-derived mesenchymal stem cells in the liver were observed by histochemistry, fluorescent labeling, and serological detection.
RESULTS: At 1, 2, and 3 weeks after adipose-derived mesenchymal stem cell injection, liver fibrosis was significantly ameliorated. The injected adipose-derived mesenchymal stem cells had hepatic differentiation potential in vivo, and the survival rate of adipose-derived mesenchymal stem cells declined over time.
CONCLUSIONS: The findings in this study confirmed that adipose-derived mesenchymal stem cells derived from the Bama pig can be used in the treatment of liver fibrosis, and the grafted adipose-derived mesenchy-mal stem cells can migrate, survive, and differentiate into hepatic cells in vivo.

Matched MeSH terms: Swine
Nipah virus infection: pathology and pathogenesis of an emerging paramyxoviral zoonosis

Wong KT, Shieh WJ, Kumar S, Norain K, Abdullah W, Guarner J, et al.

Am J Pathol, 2002 Dec;161(6):2153-67.
PMID: 12466131

In 1998, an outbreak of acute encephalitis with high mortality rates among pig handlers in Malaysia led to the discovery of a novel paramyxovirus named Nipah virus. A multidisciplinary investigation that included epidemiology, microbiology, molecular biology, and pathology was pivotal in the discovery of this new human infection. Clinical and autopsy findings were derived from a series of 32 fatal human cases of Nipah virus infection. Diagnosis was established in all cases by a combination of immunohistochemistry (IHC) and serology. Routine histological stains, IHC, and electron microscopy were used to examine autopsy tissues. The main histopathological findings included a systemic vasculitis with extensive thrombosis and parenchymal necrosis, particularly in the central nervous system. Endothelial cell damage, necrosis, and syncytial giant cell formation were seen in affected vessels. Characteristic viral inclusions were seen by light and electron microscopy. IHC analysis showed widespread presence of Nipah virus antigens in endothelial and smooth muscle cells of blood vessels. Abundant viral antigens were also seen in various parenchymal cells, particularly in neurons. Infection of endothelial cells and neurons as well as vasculitis and thrombosis seem to be critical to the pathogenesis of this new human disease.

Matched MeSH terms: Swine
Fulltext Predifferentiated amniotic fluid mesenchymal stem cells enhance lung alveolar epithelium regeneration and reverse elastase-induced pulmonary emphysema

Lan YW, Yang JC, Yen CC, Huang TT, Chen YC, Chen HL, et al.

Stem Cell Res Ther, 2019 06 13;10(1):163.
PMID: 31196196 DOI: 10.1186/s13287-019-1282-1

INTRODUCTION: Pulmonary emphysema is a major component of chronic obstructive pulmonary disease (COPD). Emphysema progression attributed not only to alveolar structure loss and pulmonary regeneration impairment, but also to excessive inflammatory response, proteolytic and anti-proteolytic activity imbalance, lung epithelial cells apoptosis, and abnormal lung remodeling. To ameliorate lung damage with higher efficiency in lung tissue engineering and cell therapy, pre-differentiating graft cells into more restricted cell types before transplantation could enhance their ability to anatomically and functionally integrate into damaged lung. In this study, we aimed to evaluate the regenerative and repair ability of lung alveolar epithelium in emphysema model by using lung epithelial progenitors which pre-differentiated from amniotic fluid mesenchymal stem cells (AFMSCs).
METHODS: Pre-differentiation of eGFP-expressing AFMSCs to lung epithelial progenitor-like cells (LEPLCs) was established under a modified small airway growth media (mSAGM) for 7-day induction. Pre-differentiated AFMSCs were intratracheally injected into porcine pancreatic elastase (PPE)-induced emphysema mice at day 14, and then inflammatory-, fibrotic-, and emphysema-related indices and pathological changes were assessed at 6 weeks after PPE administration.
RESULTS: An optimal LEPLCs pre-differentiation condition has been achieved, which resulted in a yield of approximately 20% lung epithelial progenitors-like cells from AFMSCs in a 7-day period. In PPE-induced emphysema mice, transplantation of LEPLCs significantly improved regeneration of lung tissues through integrating into the lung alveolar structure, relieved airway inflammation, increased expression of growth factors such as vascular endothelial growth factor (VEGF), and reduced matrix metalloproteinases and lung remodeling factors when compared with mice injected with AFMSCs. Histopathologic examination observed a significant amelioration in DNA damage in alveolar cells, detected by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling (TUNEL), the mean linear intercept, and the collagen deposition in the LEPLC-transplanted groups.
CONCLUSION: Transplantation of predifferentiated AFMSCs through intratracheal injection showed better alveolar regeneration and reverse elastase-induced pulmonary emphysema in PPE-induced pulmonary emphysema mice.

Matched MeSH terms: Swine
Fulltext Highly pathogenic avian influenza virus nucleoprotein interacts with TREX complex adaptor protein Aly/REF

Balasubramaniam VR, Hong Wai T, Ario Tejo B, Omar AR, Syed Hassan S

PLoS One, 2013;8(9):e72429.
PMID: 24073193 DOI: 10.1371/journal.pone.0072429

We constructed a novel chicken (Gallus gallus) lung cDNA library fused inside yeast acting domain vector (pGADT7). Using yeast two-hybrid screening with highly pathogenic avian influenza (HPAI) nucleoprotein (NP) from the strain (A/chicken/Malaysia/5858/2004(H5N1)) as bait, and the Gallus gallus lung cDNA library as prey, a novel interaction between the Gallus gallus cellular RNA export adaptor protein Aly/REF and the viral NP was identified. This interaction was confirmed and validated with mammalian two hybrid studies and co-immunoprecipitation assay. Cellular localization studies using confocal microscopy showed that NP and Aly/REF co-localize primarily in the nucleus. Further investigations by mammalian two hybrid studies into the binding of NP of other subtypes of influenza virus such as the swine A/New Jersey/1976/H1N1 and pandemic A/Malaysia/854/2009(H1N1) to human Aly/REF, also showed that the NP of these viruses interacts with human Aly/REF. Our findings are also supported by docking studies which showed tight and favorable binding between H5N1 NP and human Aly/REF, using crystal structures from Protein Data Bank. siRNA knockdown of Aly/REF had little effect on the export of HPAI NP and other viral RNA as it showed no significant reduction in virus titer. However, UAP56, another component of the TREX complex, which recruits Aly/REF to mRNA was found to interact even better with H5N1 NP through molecular docking studies. Both these proteins also co-localizes in the nucleus at early infection similar to Aly/REF. Intriguingly, knockdown of UAP56 in A549 infected cells shows significant reduction in viral titer (close to 10 fold reduction). Conclusively, our study have opened new avenues for research of other cellular RNA export adaptors crucial in aiding viral RNA export such as the SRSF3, 9G8 and ASF/SF2 that may play role in influenza virus RNA nucleocytoplasmic transport.

Matched MeSH terms: Swine

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