Plasma volume, cell volume, total blood volume and F factor in the tree shrew.

In this study, the physiological values of volumes of plasma, cells, total blood and the F blood factors were identified in 24 adult tree shrews (Tupaia belangeri; 12 male and 12 female; average BW of 123.9±19.19 g). The two-compartment model method of Evans Blue dye was used to obtain the plasma volume and the venous hematocrit was measured by microhematocrit method. To establish the relationship between body weight (BW) and blood volume of tree shrews, We performed linear fitting for these two datasets. Results were analyzed according to gender and weight (<120g vs.>120g). Statistical significance was assessed using the unpaired student t test and one-way ANOVA. The average volumes per 100g body weight of plasma, red blood cell (RBC) and total blood were 5.42±0.543, 3.24±0.445, and 8.66±0.680ml respectively. The mean body hematocrit, cardiac hematocrit, jugular vein hematocrit, femoral vein hematocrit, and tail vein hematocrit was 37.43±4.096, 39.72±3.219, 43.04±4.717, 40.84±3.041, and 38.71±3.442% respectively. The F cardiac was 0.94±0.072, F jugular vein 0.88±0.118, F femoral vein 0.92±0.111, and the F tail vein 0.97±0.117. Blood volume (ml) was 85.89103×BW (kg). This is the first study to provide the parameters of plasma volume, cell volume, total blood volume and F factor and a baseline for future research on blood physiology of tree shrews.

Development of a tree shrew-specific interferon-gamma assay.

Tree shrews (Tupaia belangeri) are small squirrel-like mammals closely related to primates. Due to their susceptibility to several human viruses, tree shrews have been proposed as potential animal models for the study of human viral infections. However, there are no standardized assays currently available for the detection of tree shrew-specific interferon (IFN)-γ, a major cytokine secreted during the antiviral immune response. Herein, we developed a novel enzyme-linked immunosorbent assay (ELISA) for the quantification of IFN-γ in tree shrew serum samples. Tree shrew-specific IFN-γ was expressed in Escherichia coli via fusion with glutathione S-transferase (GST-TS-IFN-γ) to obtain recombinant IFN-γ. To generate anti-IFN-γ monoclonal antibodies, mice were immunized with the GST-TS-IFN-γ recombinant fusion protein, and hybridoma cell lines were established. Similarly, anti-IFN-γ polyclonal antibodies were obtained from immunized rabbits, purified, and conjugated to horseradish peroxidase (HRP). Based on the results obtained from the antibody matching test, we optimized the monoclonal antibody (1:2000) and the HRP-conjugated polyclonal antibody (1:8000) as coating and detection antibodies, respectively. Titration curves were generated with recombinant IFN-γ to develop a sensitive sandwich ELISA; the lowest detection limit of the assay was 20 ng/mL. We also tested mitogen-stimulated tree shrew blood samples in this ELISA, and found significantly higher levels of IFN-γ in the stimulated versus the unstimulated samples. Most importantly, our ELISA system detected native IFN-γ in serum samples from 50 healthy tree shrews. We have thus developed a novel ELISA, and have demonstrated the first ELISA-based measurement of IFN-γ in tree shrew serum samples.

Comparative analysis of glucuronidation of ethanol in treeshrews, rats and humans.

1. The treeshrews consume food-derived alcohol (ethanol) at a dose that would intoxicate humans, highlighting a marked difference in detoxification of ethanol between the animal species and humans. 2. In this study, we reported that the treeshrews and rats exhibited considerably high glucuronidation capacity for ethanol. Ethanol glucuronidation was 7.1-fold (for the liver microsomes) or 29.2-fold (for the intestine microsomes) more efficient in treeshrews than in humans. Similar to treeshrews, rats also showed a high efficiency in glucuronidating ethanol. 3. In the single-pass perfused intestinal model, significant amount of ethyl glucuronide (EtG) was excreted into the perfusate (for both treeshrews and rats) and bile (for rats). Biliary excretion of EtG was 8.8-13.4 times of intestinal excretion of EtG, suggesting that the liver played a determinant role in glucuronidation of ethanol. In vivo pharmacokinetics showed that EtG production was rapid in the animals with a Tmax value of ≤ 1.75 h. The excreted EtG into urine was 0.11-0.13% of dosed ethanol, a value increased by a 5.5- to 6.6-fold compared to that in humans. 4. This was the first report that the glucuronidation activity toward ethanol was much higher in treeshrews and rats than that in humans, revealing a marked species difference in ethanol glucuronidation.

[Detection and comparison of physiological indexes in the wild and laboratory tree shrew].

To provide fundamental basis for the tree shrew models of human diseases, we examined and compared the physiological and biochemical indexes between wild and laboratory tree shrews. Blood samples were taken from 54 wild tree shrews that were housed in laboratory for 1-2 months, and from 54 first-generation of the laboratory tree shrews; each group had nearly equal male and female composition. Some of the first reported physiological and biochemical indexes were showed no significant differences between genders, and these indexes in laboratory tree shrews were as follows [medium (inter-quartile range) ]: CK 1449 (956) U/L, CTNI 5.94 (7.23) ug/L, TBA15.6 (19.7) µmol/L, FRUC 393.5 (80.8) µmol/L and LDL-C0.36 (0.32); and in the wild tree shrews, 986 (564) U/L, 4.01 (4.10) µg/L, 20.0 (20.6) µmol/L, 379.0 (104.0) µmol/L and 0.46 (0.23) mmol/L, respectively. In the laboratory tree shrews, the variations of physiological and biochemical indexes were smaller, but the mean values of some indicators related to liver and heart functions became higher. These data would be valuable for the development of tree shrew models of human diseases.

[Preliminary investigation of viruses to the wild tree shrews (Tupaia belangeri Chinese)].

Virological testing and monitoring is a fundamental part of quality control of experimental animals. However, there are few papers regarding the spectrum and status of natural infection in wild tree shrews with human and animal pathogenic viruses. Using enzyme-linked immunosorbent adsorption assay (ELISA), we tested sixty wild tree shrews captured from Qinglong, an outskirt region of Kunming, Yunnan Province, China for eleven viruses, including herpes simplex virus, coxsackie virus, influenza virus, HAV, HBV, HCV, HDV, dengue virus, hemorrhagic fever virus and measles virus. Our results showed that, in the serum samples, 22/60 (36.7%) and 1/60 (1.67%) were antibody positive for herpes simplex virus and coxsackie virus, respectively, and 4/60 (6.7%) were antigen positive for rotavirus in the feces. The remaining species of viruses were negative in these tree shrews. Based on these results, we propose that herpes simplex virus, coxsackie virus and cotavirus should be listed as top priority for routine virological monitoring of tree shrews.

Amino acid sequences of the alpha and beta chains of adult hemoglobin of the European hedgehog, Erinaceus europaeus.

Globin prepared from hemoglobin of the European hedgehog (Erinaceus europaeus) was separated into alpha and beta polypeptide chains by chromatography on a CM 52 column. The S-aminoethylated alpha and beta chains were each digested with trypsin and the resulting peptides were isolated. The sequences of all the tryptic peptides were established. The ordering of these peptides in the alpha and beta chains was deduced from their homology with the primary structures of alpha and beta chains of human adult hemoglobin. Comparing the primary structures of the alpha and beta chains of adult hemoglobin of the European hedgehog thus obtained with those of adult hemoglobin of the tupai (Tupaia glis), 35 amino acids substitutions in the alpha chains and 30 in the beta chains were recognized.

Amino acid sequences of the aplpha and beta chains of adult hemoglobin of the tupai, Tupaia glis.

Globin prepared from hemoglobin of adult tupai (Tupaia glis) was separated into alpha and beta polypeptide chains by CM-cellulose column chromatography. The S-aminoethylated alpha polypeptide chain and S-carboxymethylated beta polypeptide chain were each digested with trypsin, and the sequences of all the peptides thus obtained were established. The ordering of these tryptic peptides in the alpha and beta polypeptide chains was deduced from the homology of their primary structures with that of human adult hemoglobin. In this way the primary structures of the alpha and beta polypeptide chains of tupai hemoglobin were established; 27 amino acids in the alpha polypeptide chain and 26 in the beta chain differ from those in human adult hemoglobin.