Wasp-stung rat model translationally expresses the coagulopathy manifestations of human wasp patients.

Two over 80 wasp stings male victims appeared severe abnormal coagulation were consecutively examined by thromboelastography (TEG) guided with heparinase during hospitalization. However, the cause of coagulopathy remains unsolved. Rats were applied to establish a wasp-stung animal model highly resembled the manifestations of wasp-stung patients. According body surface area conversion, Sprague-Dawley rats were stung based on wasp sting numbers (0, 4, 8, 12 stings; n = 6 each) with various exposure times (0, 1, 3, 6 h) to determine the simulation of coagulopathy. The blood R, K values, and angle degree of wasp-stung rats were measured by TEG. The TEG profiles of stung rats were found to be concomitant with that of wasp-stung patients. Data showed that the endogenous heparinization of rats was time-dependent. Compared to the TEG profile of eight stings given rat, the coagulation time of 2 mm clot formation at 3 h (R value) was longer than that at 0 h. The coagulation time was prolonged with increasing sting numbers when compared to the various stings at 1, 3, and 6 h exposed. Interestingly, there was observed the peak coagulation at 3 h of eight stings. The Ck-standard and Ck-heparinase at 3 h after 8 stings given were R: 9.6-4.4 min; K: 3.8-1.8 min; angle degree: 49.8-68.0, respectively. The original data of R, K values and angle degree in two wasp-stung victims were 11.7-13.6 min, 4.3-5.5 min, and 41.2-32.8° in CK-standard, respectively; whereas those of the CK-heparinase groups were 5.6-6.7 min, 2.4-2.5 min, and 59.5-58.8°, correspondingly. Conclusively, this massive wasp-stung animal model can be applied to the investigations of pathogenesis and provides a clinical strategy or guideline for clinical intervention.

Effect of Oyster Meat Preload on Postmeal Glycemic Control in Healthy Young Adults.

Objective: Evidence suggests that food preload improves postmeal glycemic profiles, but the effects of marine food are poorly understood. Our study aims to verify the regulating effects of premeal oyster meat (OM) on postprandial blood glucose.Method: Edible parts of the flesh of oyster were prepared for a randomized crossover experiment. After overnight fasting, 20 healthy young men consumed 300 mL of preload drinks with 0 g/kg body weight (BW) (control), 0.1 g/kg BW, and 0.2 g/kg BW. Peripheral blood concentrations of glucose and gastrointestinal hormones were measured before preloading at baseline (0 minutes) and at intervals after the preload and after a preset rice meal. The nutrient composition of OM was analyzed.Results: Compared with other doses, 0.2 g/kg BW OM preload induced higher plasma premeal insulin (p < 0.05), C-peptide (p < 0.05), and glucagon-like peptide-1 (GLP-1; p < 0.05) without altering the glucose concentrations during premeal times. By contrast, 0.2 g/kg BW OM induced less secretion of glucose (p < 0.05) and gastric inhibitory peptide (GIP; p < 0.05), but higher secretion of GLP-1 (p < 0.05) than 0.1 g/kg BW of OM after a meal. During the entire experiment (0-170 minutes), OM reduced the blood glucose (p < 0.05) and GIP (p < 0.05), but increased GLP-1 (p < 0.05). OM was rich in protein (78.4%) and low in fat (6%). Glutamic acid, aspartic acids, glycine, and taurine are the amino acids with high content found in OM.Conclusions: OM preload reduces postmeal glycemia in healthy young people with associated changes in gastrointestinal hormone responses. This effect may be attributed to the rich contents of protein and amino acids of OM.

An improved gold nanoparticle probe-based assay for HCV core antigen ultrasensitive detection.

A gold nanoparticle probe-based assay (GNPA) was developed for ultrasensitive detection of Hepatitis C virus (HCV) core antigen. In the GNPA, after anti-HCV core antigen polyclonal antibodies and single-stranded barcode signal DNA were labeled on gold nanoparticle probe (NP), DNA enzyme was used to degrade the unbound barcode DNAs. The anti-HCV core antigen monoclonal antibodies were coated on magnetic microparticles probe (MMP). Then the NP-HCV core antigen-MMP sandwich immuno-complex was formed when the target antigen protein was added and captured. Magnetically separated, the immuno-complex containing the single-stranded barcode signal DNA was characterized by TaqMan probe based real-time fluorescence PCR. A detection limit of 1 fg/ml was determined for the HCV core antigen which is magnitude greater than that of ELISA (2ng/ml). The coefficients of variation (CV) of intra-assay and inter-assay respectively ranged from 0.22-2.62% and 1.92-3.01%. The improved GNPA decreased the interference of unbound barcode DNAs and may be an new way for HCV core antigen detection.

Establishment of evanescent wave fiber-optic immunosensor method for detection bluetongue virus.

The evanescent wave fiber immunosensors (EWFI) technique was developed for the real-time rapidly sensitive and specific detection of the monoclonal antibody 3E2 of BTV. The outer-core protein VP7 of BTV was labled on the surface of the exposed fiber-optic core. The monoclonal antibody 3E2 of BTV VP7 were added and then the goat ant-rat IgG conjugated with Cy3 was captured. After the 532nm pulse (excitation source) reached the fiber probe, evanescent wave was generated, which excited the Cy3 bound to the immuno-complex and produced the fluorescent signal, which was changed into electrical signals read through computer. The preliminary results suggested that a detection limit of 10ng/ml was measured for the monoclonal antibody 3E2, which is equal to the sensitivity of ELISA. The 3E2 sample was specifically detected through the EWFI assay in 15min, and the fiber can be recycled at least ten times through TEA solution condition. This developed EWFI was a real-time rapidly sensitive and specific way for the detection of BTV antibodies.

[Effects of electroacupuncture of "Futu" (LI 18), etc. on pain behavior and expression of GABA receptor subunit genes in cervical spinal cord in rats with thyroid regional pain].

OBJECTIVE: To observe the effect of electroacupuncture (EA) at different acupoints on the expression of gamma aminobutyric (GABA) receptor (R) subunit genes in the cervical spinal cord in rats with thyroid regional inflammatory pain so as to analyze its analgesic mechanism for thyroid surgery. METHODS: A total of 50 Wistar rats were randomized into control, model, Futu (LI 18), Hegu (LI 4)-Neiguan (PC 6, LI 4-PC 6), Zusanli (ST 36)-Yanglingquan (GB 34, ST 36-GB 34) groups, with 10 rats in each group. Thyroid regional pain model was established by subcutaneous injection of 2.5% formalin (100 microL). Ten minutes after modeling, EA (2 Hz/15 Hz, 1 mA) was applied to LI 18, LI 4-PC 6 and ST 36-GB 34 for 30 min, respectively. The animals' face-grooming (FG) times in 5 min and thermal pain threshold (paw withdrawal latency, PWL) were recorded. The expression of GABA(A) R, GABA(B) R1, and GABA(B) R2 genes in the cervical 1-3 segments of the spinal cord 80 min after modeling was detected by using reversed transcription-polymerase chain reaction (RT-PCR). Histological changes of the tissues of the thyroid region were observed by using H. E. staining. RESULTS: After subcutaneous injection of formalin, the animals' FG times in 5 min were increased considerably and the thermal pain threshold was decreased obviously in the model group (P < 0.05). Concomitantly, the expression levels of GABA(B) R1, and GABA(B) R2 genes of the cervical spinal cord were upregulated significantly in the model group (P < 0.05), and GABA(A) R expression was increased slightly (P > 0.05). Compared with the model group, the FG times in 5 min in the LI 18 and LI 4-PC 6 groups at 40 min and 70 min after modeling were decreased significantly (P < 0.05), and their thermal pain threshold values were increased markedly (P < 0.05). The expression levels of GABA(B) R1 mRNA, GABA(B) R2 mRNA and GABAA R mRNA were significantly higher in the LI 18 and LI 4-PC 6 groups than in the model group (P < 0.05). The expression level of GABA(B) R1 mRNA of the ST 36-GB 34 group was obviously higher than that of the model group (P < 0.05). The inflammatory cells in the formalin-injected thyroid region were relatively fewer in the LI 18 and LI 4-PC 6 groups than in the model group. CONCLUSION: Both EA of LI 18 and LI 4-PC 6 can significantly suppress formalin-injection induced pain reactions in the thyroid region, which may be closely associated with its effects in upregulating expression levels of cervico-spinal GABA(B) R1 mRNA, GABA(B) R2 mRNA and GABA(A) R mRNA, and reduce regional inflammatory reactions in the thyroid region.

[Neuroanatomical basis of clinical joint application of "Jinggu" (BL 64, a source-acupoint) and "Dazhong" (KI 4, a Luo-acupoint) in the rat: a double-labeling study of cholera toxin subunit B conjugated with Alexa Fluor 488 and 594].

OBJECTIVE: To study the specific correlation between "Jinggu" (BL 64) and "Dazhong" (KI 4) in the nervous system by using a double-labeling of cholera toxin subunit B conjugated with Alexa Fluor 488 and 594 (CTB-Alexa 488, 594) in rats, so as to investigate its neuroanatomical basis for clinical joint-application of Yuan-Source and Luo acupoints. METHODS: Three male SD rats were used in the present study. Under anesthesia (10% urethane), 0.1% CTB-Alexa 488 (5 microL) and CTB-Alexa 594 (5 microL) were respectively injected into the border area between the red and white flesh, distal to the tuberosity of the fifth metatarsal bone, and the depression anterior to the medial attachment of the calcaneal tendon, the corresponding sites of the acupoints Jinggu (BL 64) and Dazhong (KI 4) in the human body. After 3 surviving days, the rat's brain, spinal cord and dorsal root ganglia (DRGs) of L3-L6 were dissected following perfusion with 4% paraformaldehyde, cut into sections and observed under fluorescent microscope equipped with a digital camera. The labeled neurons were recorded and counted. RESULTS: It was found under fluorescent microscope that the single-labeled neurons and the dual-labeled neurons were ipsilaterally located on the injected side. Among the single-labeled neurons, the labeled sensory neurons related to "Jinggu" (BL 64) and "Dazhong" (KI 4) were found to be in the DRGs of L3-L6, with a higher concentration in the DRGs of L.4 (27/162, 102/332) and L5 (130/162, 204/332). The dual-labeled 7 neurons were found to be in DRGs of L4 and L5. In addition, the labeled motoneurons related to "Jinggu" (BL 64) and "Dazhong" (KI 4) distributed in the dorsolateral portion of lamina IX, forming a longitudianal column from L3-L6 with a higher concentration at L4 and L5. CONCLUSION: The labeled sensory and motor neurons innervating Yuan-acupoint "Jinggu" (BL 64) and Luo-acupoint "Dazhong" (KI 4) distribute in DRGs of the same spinal segments and spinal ventral horns from L3-L6.

[Effect of acupuncture of "Zusanli"(ST 36)on sexual hormone levels in spleen-deficiency syndrome rats].

OBJECTIVE: To study the underlying mechanism of acupuncture of "Zusanli" (ST 36) in clinical treatment of spleen deficiency syndrome (SDS). METHODS: Twenty-six adult male Wistar rats were randomized into normal control group (n=6), model group (n=8), saline + model (saline) group (n=6) and acupuncture + model (acupuncture) group (n=6). The spleen deficiency syndrome model was duplicated by intragastric perfusion of 50% alcohol (1 mL/100 g, once) and vinegar (pH 3, 1 mL/100 g, once daily for 10 d). Bilateral "Zusanli" (ST 36) was punctured with filiform needles which were manipulated for 1 min after insertion and retained for 20 min. The treatment was conducted once daily for 10 days. The contents of serum testosterone(T) and estradiol(E2)were detected by radioimmunoassay. The rats' physical strength was detected by exhausted swimming test in cool water. RESULTS: In comparison with the normal group, the duration of exhausted swimming, serum T and E2 contents and T/E, were decreased significantly in the model group (P < 0.05). Compared with the model group, the duration of exhausted swimming, serum T and E2 contents and T/E2 in the acupuncture group were upregulated significantly (P < 0.05). No significant differences were found between the saline and control groups (P > 0.05). CONCLUSION: Acupuncture of "Zusanli" (ST 36) is able to reverse SDS-induced decrease of sexual hormones in spleen deficiency syndrome rats, which may contribute to its effect in improving physical strength.

[Effects of electroacupuncture at "Futu" (LI 18), etc. on expression of spinal 5-HT 1 AR mRNA, 5-HT 2 AR mRNA and protein in rats with neck incision pain].

OBJECTIVE: To observe the effect of electroacupuncture (EA) at bilateral "Futu" (LI 18), etc. on the expression of 5-HT 1 A receptor (R) mRNA, 5-HT2 AR mRNA and protein in the spinal cord of rats with neck incision pain, so as to explore its underlying mechanism in relieving incision pain. METHODS: A total of 48 Wistar rats were randomly divided into control, model (incision pain), Futu (LI 18), Hegu (LI 4)-Neiguan (PC 6, LI 4-PC 6), Zusanli (ST 36)-Yanglingquan (GB 34) cervical cord (ST 36-GB 34 C) and lumbar cord (ST 36-GB 34 L) groups. A 1.5 cm longitudinal incision was made in the middle of the neck under Isoflurane inhalation anesthesia. Pain threshold (PT) was measured using radiant heat. EA (1-2 mA, 2 Hz/15 Hz) was applied to bilateral LI 18, LI 4-PC 6, ST 36-GB 34 for 30 min. The expression of 5-HT 1 AR mRNA, 5-HT 2 AR mRNA and protein in the cervical spinal cord (C1-C4) tissue, and 5-HT 1 AR mRNA and 5-HT 2 AR mRNA in the lumbar cord (L1- L3) were detected with Real-time PCR and Western blot analysis, respectively. RESULTS: After the operation, the thermal PT was shortened obviously In comparison with pre-operation (P < 0.05). Compared with post-operation, the PT values were increased markedly in the LI 18 group and LI 4-PC 6 group (P < 0.05), rather than in the ST 36-GB 34 group (P > 0.05). The expression levels of 5-HT 1 AR mRNA, 5-HT 2 AR mRNA and 5-HT 2 AR protein in the cervical cord of the model group were increased significantly compared with those of the control group (P < 0.05). In comparison with the model group, the expression level of 5-HT 1 AR mRNA was down-regulated considerably in the LI 18 and LI 4-PC 6 groups (P < 0.05), and those of spinal 5-HT 2 AR mRNA and protein were up-regulated significantly in the LI 18 and LI 4-PC 6 groups (P < 0.05). No significant changes were found in the expression of 5-HT 1 AR mRNA and 5-HT 2 AR mRNA in the ST 36-GB 34 C group in comparison with the model group (P > 0.05). The expression levels of 5-HT 1 AR mRNA and 5-HT 2 AR mRNA in the ST 36-GB 34 L group were significantly lower than those of the ST 36-GB 34 C group (P < 0.05). CONCLUSION: EA of LI 18 and LI 4-PC 6 can significantly suppress pain reaction of neck incision in the rat, which is closely associated with its effects in down-regulating the expression of 5-HT 1 AR mRNA and in up-regulating 5-HT 2 AR mRNA and 5-HT 2 AR protein in the cervical spinal cord.

[Distribution of the activated acupoints after acute gastric mucosal injury in the rat].

OBJECTIVE: To observe the dynamic distribution of the extravasated Evans Blue (EB) dye points (neurogenic inflammatory response) at the skin after acute gastric mucosal injury (AGMI) and its relation to the related regular acupoints in the locations in rats. METHODS: A total of 70 Wistar rats were randomized into normal control (n=10), normal saline (n=10), and AGMI (n=50) groups. The AGMI group was further divided into 5 h, 2 d, 3 d, 4 d and 5 d subgroups with 10 rats in each. AGMI model was duplicated by intragastric perfusion of diluted hydrochloric acid (HCl, 0.5 mol/L). Evans Blue Dye (50 mg/kg, 50 mg/mL in 0.9% saline) was given to the rats before AGMI modeling. The plasma extravasated EB points at the skin of the whole body were observed after removal of the hair. RESULTS: The extravasated EB points presented a nerve-segmental distribution, with the proportion of the points in the location being 47.5% for "Geshu" (BL 17),58. 82% for "Jizhong" (GV 6), 88.23% for "Pishu" (BL 20), 82.35% for "Weishu" (BL21), 17.64% for "Zhongwan" (CV 12), and 5.88% for "Shangwan" (CV 13), respectively. The plasma extravasation of EB seldom appeared in normal rats and only fewer points were found in rats accepted administration of 0.9% saline. Significant differences were found between model and normal control groups, and between model and normal saline groups in the numbers of the extravasated EB points (P < 0.01, P < 0.05). The number of the extravasated EB points was related to the phase of gastric mucosa injury, being most on the 2nd and 3rd day after modeling and disappearing gradually along with the natural repair of AGMI. CONCLUSION: AGMI promotes the plasma extravasation of EB and the extravasated EB points present a nerve-segmental distribution and have a higher corresponding rate with some acupoints including "Pishu" (BL 20), "Weishu" (BL 21), etc., suggesting an activation of the normally silent acupoints under diseased conditions.

[Effect of electroacupuncture on cortical spreading depression and plasma CGRP and substance P contents in migraine rats].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on cortical spreading depression (CSD) and contents of plasma calcitonin gene-related peptide (CGRP) and substance P (SP) in migraine rats. METHODS: Thirty male SD rats were equally randomized into control, model and EA groups. Migraine model was established by topical application of KCI (3 mol/L) immersed in a piece of filter paper to the cerebral cortex (parietal lobe, 6 mm posterior to the Bregma and 5 mm to the sagital fissure) after exposure of the skull (in reference to Michael' method). KCI stimulation evoked CSD potentials (3 mm rostral to the Bregma, and 2 mm to the sagital fissure) were recorded by using a glass microelectrode. For rats of control group, filter paper containing 0.9% NaCl was applied to the same parietal cortex area. EA (1 mA, 2 Hz/100 Hz) was applied to bilateral "Yanglingquan" (GB 34) and "Taichong" (LR 3) for 30 min. The contents of plasma CGRP and SP were assayed by radioimmunoassay. RESULTS: CSD was induced 3-5 min after application of KCI to the parietal lobe. The average amplitude of model group was (-25.13 +/- 1.23) mV, and that of EA group was (-19.19 +/- 1.53) mV, displaying a significant reduction of CSD amplitude after EA (P < 0.01). Comparison among 3 groups showed that both plasma CGRP and SP contents in model group were significantly higher than those of control group (P < 0.001, P < 0.01), while compared with model group, plasma CGRP and SP levels in EA group decreased considerably (P < 0.05, P < 0.001), suggesting an inhibitory effect of EA on pain-producing substance. CONCLUSION: EA of GB 34 and LR 3 can effectively suppress KCI provoked cortical spreading depression and plasma CGRP and SP levels in the rat, which may contribute to its effect in relieving migraine in clinic.