Objective Clarify the effect of neutrophil extracellular traps (NETs) on endothelial cell injury, and investigate whether the heparin can exert a protective effect on endothelial cells by reducing the endothelial cell injury induced by NETs.Methods Neutrophils (PMN) were obtained from healthy human peripheral blood by Percoll-Histopaque density gradient centrifugation, and was stimulated with phorbol ester (PMA) to induce NETs. The qualitative and quantitative analysis of NETs was detected by immunofluorescence staining combined with fluorescence detector. The NETs were used to induce human umbilical vein endothelial cells (HUVEC)in vitro. Recombinant DNA hydrolytic enzymes (rhDNase) and heparin intervention were added respectively. The activity of HUVEC was measured by methyl thiazolyl tetrazolium (MTT) method after 6 hours.Results PMA can stimulate PMN to produce NETs. Immunofluorescence staining showed the formation of reticular formation around the PMN. The concentration of cell-free DNA in the supernatant of PMN stimulated by PMA was significant increased compared with the control group through the detection of PicoGreen fluorescent labeling instrument (2 hours: 119.62±14.83 vs. 24.27±0.67, 4 hours: 146.67±21.24 vs. 28.35±2.98, bothP < 0.05). Application of NETs to stimulate the HUVEC, cell damage was dose dependent and inhibition rate increased gradually. The endothelial cell inhibition induced by NETs can be antagonized after adding rhDNase [10μg/L NETs: (8.65±0.51)% vs. (10.99±0.35)%, 20μg/L NETs:(14.85±0.43)% vs. (16.85±0.49)%, 30μg/L NETs: (26.06±3.51)% vs. (27.54±0.62)%, allP < 0.05]. Heparin with different concentrations were added into the experimental group (0.01, 0.1, 1, 10 kU/L). We found that the endothelial cell inhibition rate decreased compared with control group [10μg/L NETs: (8.96±0.70)%, (5.32±1.36)%, (0.70±0.30)%, (0.75±0.20)% vs. (10.99±0.35)%; 20μg/L NETs: (15.57±0.62)%, (13.28±0.65)%, (6.91±0.15)%, (5.86±0.17)% vs. (16.85±0.49)%; 30μg/L NETs: (30.49±0.74)%, (29.41±1.41)%, (23.45±0.75)%, (21.72±1.52)% vs. (27.54±0.62)%, allP < 0.05].Conclusions NETs can induce endothelial cell injury, and the injury degree was increased with the concentration of NETs. Heparin can reduce endothelial cell injury induced by NETs, which may be a potential mechanism for the protective effect of heparin on sepsis.

Objective: By the bibliometrical analysis and evaluation of acupuncture theses on auricular acupoint sticking from 2005 to 2014, the analysis was processed from multiple aspects including the types and numbers of diseases and effective rates of various diseases, in order to objectively reflect the general situation of clinical application of ear point embedding method in the recent years. Methods: By the bibliometrical analysis, the literature on auricular acupoint sticking in the recent ten years was counted up, analyzed and categorized. Results and Conclusion: Auricular acupoint sticking is used clinically for diseases of 16 systems, including 130 diseases. 74 diseases were singularly treated with auricular acupoint sticking, including the advantageous disease category like insomnia, constipation and obesity, and secondary advantageous disease category like dysmenorrhea, myopia, allergic rhinitis, post-operative pain and acne vulgaris.

OBJECTIVETo explore the relationship between the needling sensation of catgut embedding therapy and the depth of embedded catgut so as to improve the safety of the needle insertion and catgut implantation of the therapy.

METHODSTwenty healthy adults were selected. Under the ultrasound, the structure of the cervical Jiaji (EX-B 2) was observed. In the ultrasound guidance, the catgut was embedded. The two-dimensional imaging method was adopted to observe the anatomic structure and the procedure of needle insertion at the cervical Jiaji (EX-B 2). The high-frequency ultrasound was used to collect the images at Jiaji (EX-B 2) of C5 and determine the depths from the skin surface to the different layers of the point. Additionally, the visual analogue scale (VAS) was adopted to score the needling sensations when the needle inserted at different layers. The persistent sensation duration in the local area was followed continuously.

RESULTSUnder the ultrasound, the anatomic structure and tissue layers of cervical Jiaji (EX-B 2) were displayed clearly. The difference was significant in the average depth from the skin surface to the subcutaneous tissue, trapezius, splenius capitis, semispinalis capitis, semipinalis cervicis, multifidus and vertebral arch between the males and females (all P<0. 01). During the needle insertion, the sensations were apparently different when the implantation went to different layers. The qi arrival presented when the catgut was embedded to the trapezius, splenius capitis, semispinalis capitis, semipinalis cervicis and multifidus. But the distending pain was the most significant when in the myofascial. Commonly, the embedded catgut 2. 5 cm in length may be implanted deeply to the multifidus and the local needling sensation lasted averagely for (72. 0 ± 10. 2) h. Conclusion Under the ultrasound guidance, the depth of embedded catgut is clearly displayed at cervical Jiaji (EX-B 2). The needle insertion and the implanted material are visible, and the relationship between qi arrival and the layer of needle insertion is determined. The accuracy and safety of minimally invasive catgut embedding therapy is improved in the treatment of cervical spondylosis.

Acupuncture Points ; Acupuncture Therapy ; Adult ; Aged ; Catgut ; utilization ; Female ; Humans ; Male ; Middle Aged ; Neck Muscles ; anatomy & histology ; diagnostic imaging ; Qi ; Sensation ; Spondylosis ; diagnostic imaging ; therapy ; Ultrasonography

Objective To investigate the influence of heparin pretreatment on serum and lung tissue level of neutrophil extracellular traps (NETs) in septic mice model and its molecular mechanism.Methods Ninety male C57BL/6J mice were randomly divided into control group (n = 30), lipopolysaccharides (LPS) group (n = 30, 30 mg/kg LPS in 100μL normal saline was intraperitoneally injected) and LPS+heparin group (n = 30, 8 U of heparin in 20μL normal saline was subcutaneously injected 30 minutes before the injection of LPS). Six hours later of LPS injection, blood was collected and lung tissue was harvested. Enzyme linked immunosorbent assay (ELISA) was used to assess the concentration of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and histones 2AX (H2AX), neutrophil elastase (NE), which reflected NETs concentration. PicoGreen fluorescent dyes was used to detect serum circulating free DNA (cf-DNA/NETs) concentration. The protein expression levels of H2AX and NE in lung tissue were examined by Western Blot.Results The serum concentrations of TNF-α, IL-6, H2AX, NE, cf-DNA/NETs, and the protein expression levels of H2AX and NE in lung tissue of septic mice were significantly higher than those of control group [TNF-α (ng/L): 133.0±14.1 vs. 2.7±1.0, IL-6 (ng/L): 3911.2±189.2 vs. 298.9±52.5, H2AX (ng/L): 545.5±40.0 vs. 21.9±8.3, NE (μg/L): 6.48±0.12 vs. 0.47±0.15, cf-DNA/NETs (μg/L): 846.3±137.5 vs. 152.7±36.4, H2AX protein (gray value): 1.14±0.09 vs. 0.68±0.04, NE protein (gray value): 0.56±0.03 vs. 0.32±0.04, allP < 0.05]. After heparin pretreatment, levels of serum TNF-α, H2AX, NE, cf-DNA/NETs, and protein expression levels of H2AX and NE in lung tissue were significantly reduced [TNF-α (ng/L): 83.2±7.6 vs. 133.0±14.1, H2AX (ng/L): 435.0±39.0 vs. 545.5±40.0, NE (μg/L): 4.26±0.17 vs. 6.48±0.12, cf-DNA/NETs (μg/L): 606.5±73.9 vs. 846.3±137.5, H2AX protein (gray value): 0.91±0.03 vs. 1.14±0.09, NE protein (gray value): 0.42±0.03 vs. 0.56±0.03, allP < 0.05], but no significant change was found in IL-6 (ng/L: 3919.9±166.6 vs. 3911.2±189.2,P > 0.05).Conclusion Heparin pretreatment could significantly decrease the level of NETs in serum and lung tissue, and can be the potential mechanism of its organ protection in sepsis.

Objective:To observe the effect of herbal-partitioned moxibustion (HPM) on pain-related behavior and emotion in a rat model of chronic inflammatory visceral pain, and to investigate the mechanism.
Methods:Twenty-four male Sprague-Dawley (SD) rats were randomly divided into three groups:a normal group, a model group and an HPM group. Except for the normal group, rats in the other two groups were clystered with mixed liquor of Trinitrobenzene Sulfonic Acid (TNBS) and 50%ethanol to induce the chronic inflammatory visceral pain model. After the models were established successfully, rats in the HPM group were treated with HPM at bilateral Tianshu (ST 25) and Qihai (CV 6). Rats in the normal group and the model group were only fixed as those in the HPM group without treatment. Abdominal withdrawal reflex (AWR) score, mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were adopted to detect the visceral and somatic pain;meanwhile, open field test (OFT) and elevated plus maze test (EPMT) were employed to evaluate pain emotions such as depression and anxiety.
Results:Compared with the normal group, AWR scores of the model group were significantly increased under different stimulus expansion pressure level (P<0.01), MWT and TWL were significantly decreased (P<0.05);in OFT, the values of horizontal activities and vertical activities were significantly decreased (P<0.01);in EPMT, the proportion of the number of entry into the open arms (OE%) and that of residence time in the open arms (OT%) significantly decreased (P<0.01), indicating that the model was successful. Compared with the model group, the AWR score of the HPM group was decreased significantly (P<0.05), MWT and TWL were significantly increased (P<0.05), the values of horizontal activities and vertical activities in the model group were significantly increased (P<0.01);in OFT and EPMT, OE%and OT%were significantly increased (P<0.01).
Conclusion:HPM has analgesic effect on chronic inflammatory visceral pain. It can reduce the visceral and somatic pain in rats and markedly improve the emotions such as anxiety and depression induced by chronic visceral pain.

Objective: To evaluate the therapeutic efficacy of acupuncture plus Dang Gui Bu Xue Qu Feng Tang for benign essential blepharospasm (BEB). Methods: A prospective randomized controlled trial was performed. A total of 105 participants were randomized 1:1:1 into an acupuncture group, a herbal medicine group and an acupuncture plus herbal medicine group. Participants in the acupuncture group received manual acupuncture treatment, twice a week. Participants in the herbal medicine group received Dang Gui Bu Xue Qu Feng Tang, oral administration, once a day. Participants in the acupuncture plus herbal medicine group received both treatments. The therapeutic effects of the three groups were evaluated after four weeks of treatment. The primary outcome was the Jankovic rating scale (JRS) score, and the secondary outcome was the blepharospasm disability index (BSDI) score. Results: After four weeks of treatment, the JRS total scores significantly decreased in all three groups versus baseline (P<0.05). A greater reduction in the JRS total score was reported in participants in the acupuncture plus herbal medicine group (P<0.05), but there was no significant difference between the acupuncture group and the herbal medicine group (P>0.05). The acupuncture plus herbal medicine group had a greater decrease in the JRS severity score than the herbal medicine group (P<0.05). The reduction in the JRS frequency score was not significantly different among the three groups (P>0.05). The BSDI scores significantly decreased in all three groups versus baseline (P<0.05), but the reduction in the BSDI score was insignificantly different among the three groups (P>0.05). Conclusion: It is effective in the treatment of BEB either to use acupuncture and Dang Gui Bu Xue Qu Feng Tang alone or in combination. The combination therapy shows a more significant effect than either of the treatment alone.

Objective: To observe the efficacy of electroacupuncture (EA) in treating dry eye syndromes (DES) and its regulatory effects on patient's quality of life (QOL), anxiety, and depression.Methods: By following a randomized controlled study design, eligible DES patients were allocated to an EA group (EAG) or an acupuncture group (AG). Patients in the AG were intervened by acupuncture alone, while in the EAG, electrical stimulation was applied to Cuanzhu (BL2) and Taiyang (EX-HN5) in addition to the same treatment in the AG. Changes in clinical manifestations, ocular surface disease index (OSDI), Schirmer tear test-1 (STT-1), tear-film break-up time (TF-BUT), cornea fluorescein staining (CFS), and corneal sensitivity (CS) were observed, as well as changes in the short-form 36-item health survey (SF-36) and hospital anxiety and depression scale (HADS) scores. Results: The scores of clinical symptoms, OSDI, and CFS decreased significantly in both EAG and AG when compared with the baseline values (P<0.05), respectively, while the TF-BUT and CS increased significantly (P<0.05), and the tear secretion of patients in the EAG increased (P<0.05). Compared with those before treatment, the scores of physical role functioning, general health perception, emotional role functioning, and mental health of patients in both EAG and AG showed significant increases after treatment (P<0.05), together with a marked decrease in the score of anxiety (P<0.05). Compared with the AG, the EAG obtained a more significant increase in STT-1 (P<0.05).Conclusion: DES patients respond well to both EA and acupuncture treatments, manifested by prolonged TF-BUT and improvement of CS. Meanwhile, the two approaches can improve QOL and reduce anxiety. Furthermore, EA appears more effective in increasing tear secretion than acupuncture alone.

Objective:To observethe influence of electroacupuncture (EA) on histomorphologies of lacrimal glands, cornea and conjunctiva in experimental dry eye syndrome, and to explore the repair effects of EA on lacrimal glands and ocular surface damage. Methods:Twenty-four healthy male New Zealand rabbits were randomly divided into a normal group, a model group, an EA group and a medication group, 6 rabbits in each group. Experimental dry eye syndrome models were prepared in rabbits by using 0.1% benzalkonium chloride for eye drops. Tear secretion volume, break-up time of tear film (BUT) and corneal fluorescein staining score were observed before and after the treatment. Periodic acid Schiff (PAS) staining method was used to observe the changes of conjunctival goblet cells in rabbits. After hematoxylin eosin (HE) staining, morphological changes of rabbit cornea, conjunctiva and lacrimal gland tissues were observed under light microscope. Results: Compared with the normal group, tear secretion volume and BUT were significantly reduced (bothP<0.01), while the corneal fluorescein staining score was significantly increased (P<0.01) in the model group. Compared with the model group, tear secretion volume and BUT were significantly increased, while the corneal fluorescein staining score was significantly decreased in the EA group and the medication group (allP<0.01). Compared with the normal group, the number of conjunctival goblet cells in the model group was significantly reduced; compared with the model group, the numbers of conjunctival goblet cells were all relatively higher in the EA group and the medication group. Pathological lesions of cornea, conjunctiva and lacrimal glands all showed improvement by HE staining in the EA group and the medication group after the intervention. Conclusion:EA can improve tear secretion and tear film stability of rabbit dry eye syndrome, and repair the pathologic lesions of conjunctival goblet cells, corneal epithelia, cornea, conjunctiva and lacrimal glands.

Objective To explore the clinical features and risk factors associated with intrahepatic and hilar cholangiocarcinoma after liver transplantation .Methods Retrospective analysis of clinical data was performed for 20 hospitalized patients with intrahepatic and hilar cholangiocarcinoma from June 25 ,2014 to October 31 ,2018 .Treatments and follow-up outcomes were analyzed .The survival rate was calculated by the Kaplan-Meier method and the survival curve plotted .Cox regression model was employed for analyzing the prognostic factors .Results The cumulative recurrence rate of patients with AJCC stage Ⅰ /Ⅱ was significantly lower than that in AJCC stage Ⅲ/Ⅳ .And the cumulative recurrence rate of stageⅠ/Ⅱ Patients was 0 and that of stage Ⅲ/Ⅳ 76％ (P=0 .042) .Cox regression model showed that CA19-9 was the only prognostic factor .An elevated level of CA19-9 was associated with high recurrence post-transplantation (HR=1 .001;95％ CI:1 .000～1 .001;P=0 .035) .Conclusions During progressive stage ,the recurrence rate is higher with a worse prognosis .And an elevation of CA19-9 is an independent poor prognostic factor after intrahepatic and hilar cholangiocarcinoma transplantation .

We retrospectively analyzed a child with early-onset globoid cell leukodystrophy(Krabbe's disease)caused by complex heterozygous variations in the GALC gene.The girl was admitted to the hospital at the age of 4 month with main complaints of"No obvious cause of milk refusal,poor mental state,drowsiness,convulsions,fever."Brain MRI showed abnormal symmetric signals changes in bilateral cerebellar hemispheres,bilateral internal capsule hind limbs and bilateral ventricles,thin corpus callosum,myelination process lags behind the level of children of the same age.High-throughput sequencing analysis identified compound heterozygous mutations in GALC gene(NM 000153.4):c.[908+1G>A];[194G>A and the two heterozygous mutations were correspondingly inherited from his father and mother,respectively.The application of high-throughput sequencing technology can diagnose Krabbe disease efficiently and accurately,which assists in clinical identification and diagnosis.