Objective To explore the construction of α-1,3-galactosyltransferase (GGTA1) gene-knockout (GTKO) Diannan miniature pigs and the kidney xenotransplantation from pigs to rhesus macaques, and to assess the effectiveness of GTKO pigs. Methods The GTKO Diannan miniature pigs were constructed using the CRISPR/Cas9 gene-editing system and somatic cell cloning technology. The phenotype of GTKO pigs was verified through polymerase chain reaction, Sanger sequencing and immunofluorescence staining. Flow cytometry was used to detect antigen-antibody (IgM) binding and complement-dependent cytotoxicity. Kidney xenotransplantation was performed from GTKO pigs to rhesus macaques. The humoral immunity, cellular immunity, coagulation and physiological indicators of the recipient monkeys were monitored. The function and pathological changes of the transplanted kidneys were analyzed using ultrasonography, hematoxylin-eosin staining, immunohistochemical staining and immunofluorescence staining. Results Single-guide RNA (sgRNA) targeting exon 4 of the GGTA1 gene in Diannan miniature pigs was designed. The pGL3-GGTA1-sgRNA1-GFP vector was transfected into fetal fibroblasts of Diannan miniature pigs. After puromycin selection, two cell clones, C59# and C89#, were identified as GGTA1 gene-knockout clones. These clones were expanded to form cell lines, which were used as donor cells for somatic cell nuclear transfer. The reconstructed embryos were transferred into the oviducts of trihybrid surrogate sows, resulting in 13 fetal pigs. Among them, fetuses F04 and F11 exhibited biallelic mutations in the GGTA1 gene, and F04 had a normal karyotype. Using this GTKO fetal pig for recloning and transferring the reconstructed embryos into the oviducts of trihybrid surrogate sows, seven surviving piglets were obtained, all of which did not express α-Gal epitope. The binding of IgM from the serum of rhesus monkey 20# to GTKO pig PBMC was reduced, and the survival rate of GTKO pig PBMC in the complement-dependent cytotoxicity assay was higher than that of wild-type pig. GTKO pig kidneys were harvested and perfused until completely white. After the left kidney of the recipient monkey was removed, the pig kidney was heterotopically transplanted. Following vascular anastomosis and blood flow restoration, the pig kidney rapidly turned pink without hyperacute rejection (HAR). Urine appeared in the ureter 6 minutes later, indicating successful kidney transplantation. The right kidney of the recipient was then removed. Seven days after transplantation, the transplanted kidney had good blood flow, the recipient monkey's serum creatinine level was stable, and serum potassium and cystatin C levels were effectively controlled, although they increased 10 days after transplantation. Seven days after transplantation, the levels of white blood cells, lymphocytes, monocytes and eosinophils in the recipient monkey increased, while platelet count and fibrinogen levels decreased. The activated partial thromboplastin time, thrombin time and prothrombin time remained relatively stable but later showed an upward trend. The recipient monkey survived for 10 days. At autopsy, the transplanted kidney was found to be congested, swollen and necrotic, with a small amount of IgG deposition in the renal tissue, and a large amount of IgM, complement C3c and C4d deposition, as well as CD68⁺ macrophage infiltration. Conclusions The kidneys of GTKO Diannan miniature pigs may maintain normal renal function for a certain period in rhesus macaques and effectively overcome HAR, confirming the effectiveness of GTKO pigs for xenotransplantation.

【Objective】 To investigate the resource allocation status of blood testing laboratories in 14 blood stations in Gansu Province, explore the impact of differences in basic conditions on the comprehensive testing ability of laboratories, so as to promote the homogenization and standardization of blood screening capacity in blood stations in Gansu and improve blood safety and effectivenes. 【Methods】 An evaluation index system of laboratory resource allocation was constructed and a question-naire was designed. The data of human resources, infrastructure and key equipment of 14 blood stations were collected. The entropy weight -TOPSIS method was used to evaluate and rank the resource allocation of 14 blood stations. 【Results】 In the comprehensive evaluation of blood testing laboratory resource allocation in 14 blood stations in Gansu, the top three were laboratories A, B and I, and the last three were laboratories G, M and J. On the whole, the main issue was unreasonable structure of human resources: most laboratories had unreasonable age structure; except for Laboratory A, there was no personnel with bachelor's degree or above in laboratories; most laboratories had not established a team with intermediate professional titles. In terms of infrastructure, the size of seven laboratories could not meet the needs of modern laboratory testing, and all eight blood stations had no spare nucleic acid laboratories nor a mutual spare laboratory with other blood stations As for the key equipment, 5 laboratories had no automatic blood grouping diagnostic instrument, 5 laboratories only had one set of enzyme immunoassay detection system, 3 laboratories had no spare equipment for the key equipment, which means if the equipment failure could not be repaired in time, the release of results would be affected. 【Conclusion】 There were significant differences in human resources, infrastructure and key equipment of blood testing laboratories in 14 blood stations in Gansu, which had a great impact on laboratory testing capacity and subsequent development. It is suggested that governments at all levels and health administrative departments optimize the input of laboratory resource allocation according to the blood collection volume of blood stations to gradually narrow the differences in resource distribution between different regions, improve the degree of laboratory automation and optimize the personnel structure, so as to build high-quality and efficient blood testing laboratories and ensure the safety of clinical blood use.

Here, we took base construction of neurosurgery as example to discuss and analyze according to requirements and evaluation indexes of base construction in Xinqiao Hospital, and put forward the specific objectives, measures and implementations of base construction. Foremost, we summarized experiences and overcame shortcomings through interpreting and implementing scheme of our base construction, which would help to improve the construction of standardized residency training base in China.

The adjacent anatomy of the pelvis is complicated, with digestive, urinary, reproductive and other organs as well as important blood vessels and nerves. Therefore, accurate resection of pelvic tumors and precise reconstruction of defects after resection are extremely difficult. The development of medical 3D printing technology provides new ideas for precise resection and personalized reconstruction of pelvic tumors. The “triune” application of 3D printing personalized lesion model, osteotomy guide plate and reconstruction prosthesis in pelvic tumor limb salvage reconstruction treatment has achieved good clinical results. However, the current lack of normative guidance standards such as preparation and application of 3D printing personalized lesion model, osteotomy guide plate and reconstruction prosthesis restricts its promotion and application. The formulation of this consensus provides normative guidance for 3D printing personalized pelvic tumor limb salvage reconstruction treatment.

Raspberry ketones have important therapeutic properties such as anti-influenza and prevention of diabetes. In order to obtain raspberry ketone from Chlamydomonas reinhardtii, two enzymes catalyzing the last two steps of raspberry ketone synthesis, i.e. 4-coumaryl-CoA ligase (4CL) and polyketide synthase (PKS1), were fused using a glycine-serine-glycine (GSG) tripeptide linker to construct an expression vector pChla-4CL-PKS1. The fusion gene 4CL-PKS1 driven by a PSAD promoter was transformed into a wild-type (CC125) and a cell wall-deficient C. reinhardtii (CC425) by electroporation. The results showed the recombinant C. reinhardtii strain CC125 and CC425 with 4CL-PKS1 produced raspberry ketone at a level of 6.7 μg/g (fresh weight) and 5.9 μg/g (fresh weight), respectively, both were higher than that of the native raspberry ketone producing plants (2-4 μg/g).
Acyl Coenzyme A ; Butanones ; Chlamydomonas reinhardtii/genetics* ; Ligases ; Polyketide Synthases

Objective:To provide reference for the development of a more intelligent and systematic nursing clinical decision support system based on the concept of precision nursing and data sharing, the nursing plan module of clinical decision support system.Methods:An evidence-based knowledge base was constructed based on the nursing process and the standardized nursing terminologies; the nursing plan module was designed according to clinical needs, and the logical reasoning rules were formulated from the generation, sequencing and stopping of nursing problems, objectives, measures and activities, and finally the nursing plan module of clinical decision support system was formed.Results:The nursing plan module of clinical decision support system included the basic information of patients, positive evaluation items and weight values, nursing problems, objectives, measures and activities, etc. the module could automatically deduce the nursing plan according to the patient's individual characteristic index (positive evaluation item), and sort the nursing problems and corresponding measures and activities according to the generation time, weight value and correlation degree. It could automatically distinguish nursing problems, goals, measures and the time of activity stop, and realize intelligent decision-making.Conclusion:The interface of nursing plan module of this system is clear and logical reasoning rules are rigorous. It breaks through the bottleneck of nursing decision-making based on personal professional knowledge and experience in clinical situation for a long time, which can ensure the homogeneity of nursing plan and improve the correctness of decision-making.

Objective:To explore the clinical effect of high-titer plasma in the treatment of severe viral pneumonia in children.Methods:A total of 80 eligible children with severe viral pneumonia in intensive care unit of Guangzhou Women and Children Medical Center were enrolled in this retrospective study from January 2016 to June 2019, According to whether high-titer plasma was used or not, patients were divided into the high-titer plasma group (40 cases) and non-high-titer plasma group (40 cases). Chi-square test, Fisher's exact probability test and Mann-Whitney U test were used to compare the basic data, infection indicators, blood gas and ventilator parameters related indicators, treatment and prognosis of the two groups. Results:There were no significant differences in age, sex, admission weight, onset days and fever days between the high-titer plasma group and non-high- titer plasma group before entering the study (all P>0.05); There were no significant differences in the worst SOFA score, Murray lung injury score, chest X-ray involvement and sepsis within 3 days (all P>0.05). There were no significant differences in white blood cells, neutrophils and c-reactive protein between the two groups before and after entering the study (day 3 and 7) (all P> 0.05). There were no significant differences in arterial partial pressure of oxygen (PaO 2), partial pressure of carbon dioxide (PaCO 2), lactic acid (Lac), mean airway pressure (MAP), oxygenation index (P/F) and oxygen index (OI) between the two groups before and after entering the study (day 1, 3 and 7) (all P>0.05). There were no significant differences in the use of immunoglobulin, hormone, fiberoptic bronchoscope, blood purification, high frequency ventilator, pulmonary surfactant and extracorporeal membrane between the two groups (all P>0.05). There were no statistical differences in the number of days using invasive ventilator, the number of days staying in PICU and the total length of hospital stay between the two groups (all P>0.05). There was shorter time of the body temperature decreased to normal time in the high-titer plasma group than in the non-high-titer plasma group ( Z=-2.10, P=0.04). The survival rate of the high-titer plasma group was higher than that of the non-high-titer plasma group (85% vs 65%, χ2=4.27, P=0.04). The average daily cost of the high-titer plasma group was less than that of the non-high-titer plasma group, 3 688.38 yuan (2 335.49, 5 741.51) yuan vs 3 979.24 yuan (2 670.68, 9 992.62) yuan, but the difference was not statistically significant (Z=-1.35, P=0.18). No serious adverse reactions were observed during the treatment with high-titer plasma. Conclusions:The treatment of severe viral pneumonia with high-titer plasma can shorten fever time and improve survival rate. High-titer plasma can be used as a safe and effective treatment for severe viral pneumonia.

Objective:To explore more clinical characteristics and outcomes of children with severe pneumonia induced by adenovirus type 7(Adv-7).Methods:Seventy children with severe pneumonia induced by Adv-7 admitted to the pediatric intensive care unit of Guangzhou Women and Children′s Medical Center from January 2016 to October 2019 were enrolled.The clinical features, treatments and outcomes of these children were recorded.Results:(1)There were 43 males(61.4%)and 27 females(38.6%). Thirty(42.9%)patients were 0-12 months old, 28(40.0%)patients were 13-36 months old, and 12(17.1%)patients were over 36 months old.(2)The average onset time of severe pneumonia induced by Adv-7 was(11.87±7.10)days before being admitted to PICU.The average sequential organ failure assessment score was 6.80±3.13.The average Murray lung injury score was 2.49±1.15.The average P/F value was(150.57±86.25)mmHg(1 mmHg=0.133 kPa). Sixty-four cases(91.4%)were involved in two or more area of lung on X-rays.All cases were diagnosed with sepsis.(3)Laboratory examination: white blood cell count was(7.6±5.5) ×10 9/L, platelet count was(238.8±164.2)×10 9/L, and C-reactive protein was(39.4±37.2)mg/L.(4)Treatments: intravenous gamma globulin was used in 65 cases(92.9%). Intravenous glucocorticoid was used in 45 cases(64.3%). Fiberoptic bronchoscopy was performed in 43 cases(61.4%). Blood purification treatment was performed in 21 cases(30%). Sixty-three patients(90.0%)were treated with non-invasive or invasive ventilator, high frequency ventilator-assisted ventilation was used in 20 cases(28.6%). Surfactant was used in six cases(8.6%). Extracorporeal membrane oxygenation was used in 19 cases(27.1%). (5)The average treatment time for ventilation was(13.10±11.58)days.The average time for temperature recover was(4.69±4.01)days in PICU.The average PICU stay time was(15.76±12.20)days.The average length of stay was(27.04±13.10)days.There were 16 patients died and the mortality was 22.9%. Conclusion:The children with severe pneumonia induced by Adv-7 were critical and had significant lung injuries.Suffering from active clinical treatment such as extracorporeal membrane oxygenation, some of these patients still had poor prognosis.

Objective:To explore the efficacy and safety of high-titer plasma in the treatment of pediatric patients with severe adenovirus pneumonia.Methods:The clinical data of 92 pediatric patients with severe adenovirus pneumonia admitted to pediatric intensive care unit (PICU) in Guangzhou Women and Children′s Medical Center from January 2016 to October 2019 were retrospectively collected. According to the treatment with or without high-titer plasma, the patients were divided into plasma treatment group ( n=41) and non-plasma treatment group ( n=51). The 51 patients with chest radiograph showing more than half the lungs involved were divided into plasma treatment group ( n=29) and non-plasma treatment group ( n=22). According to fever duration before plasma treatment, patients were divided into early group (≤5 days, n=5), middle group (>5-10 days, n=14), and late group (>10 days, n=22). Baseline data, therapeutic effects, and prognosis of patients in each group were analyzed with t test, non-parametric rank sum test, one-way ANOVA and chi-square test. Results:Ninety-two patients were included. There were no significant differences in age, gender, body weight, fever duration, sequential organ failure assessment, and Murray lung injury score between plasma treatment group and non-plasma treatment group before admission (all P>0.05). The proportion of patients whose temperature drop to normal within 5 days was higher in plasma treatment group than that in non-plasma treatment group (88% (36/41) vs. 69% (35/51), χ 2=4.745, P=0.029). However, there were no significant differences between the two groups in the proportions of invasive ventilator weaning within 14 days (63% (26/41) vs. 76% (39/51), χ 2=1.868, P=0.172), transfer out from PICU within 14 days (49% (20/41) vs. 69% (35/51), χ 2=3.724, P=0.054), discharge within 28 days (51% (21/41) vs. 61%(31/51), χ 2=0.846, P=0.358) and survived patients (85% (35/41) vs. 76%(39/51), χ 2=1.143, P=0.285). Among patients with severe chest radiograph, the proportions of patients whose temperature drop to normal within 5 days and survived patients were higher in plasma treatment group than those in non-plasma treatment group (86% (25/29) vs. 59% (13/22), χ 2=4.843, P=0.028; 83% (24/29) vs. 55%(12/22), χ 2=4.796, P=0.029, respectively). However, there were no significant differences between the two groups in the proportions of invasive ventilator weaning within 14 days (52% (15/29) vs. 59% (13/22), χ 2=0.274, P=0.601), transfer out from PICU within 14 days (34% (10/29) vs. 45% (10/22), χ 2=0.632, P=0.427), and discharge within 28 days (45% (13/29) vs. 45% (10/22), χ 2=0.002, P=0.964). Among early, middle and late group, the proportions of invasive ventilator weaning within 14 days were 2/5, 13/14 and 50% (11/22), respectively, with statistically significant difference (χ 2=8.119, P=0.017). There were no significant differences in the proportions of patients whose temperature drop to normal within 5 days (4/5, 14/14, 82% (18/22), χ 2=2.965, P=0.227), transfer out from PICU within 14 days (2/5, 10/14, 36%(8/22), χ 2=4.386, P=0.112), discharge within 28 days (2/5, 8/14, 50% (11/22), χ 2=0.462, P=0.794) and survived patients (4/5, 13/14, 82% (18/22), χ 2=0.966, P=0.617) in the three groups. Only one case with high-titer plasma therapy had rash in the course of infusing plasma and no other adverse reactions were observed. Conclusions:High-titer plasma can shorten the fever time and improve the proportion of survival patients in pediatric severe adenovirus pneumonia. The clinical effect of high-titer plasma is better in 5-10 days of fever course. High-titer plasma is an effective and safe treatment.

Objective To compare the hemodynamic characteristics in internal carotid artery models, which were obtained by multi-scale unidirectional and bidirectional coupling models, so as to provide references for selecting models in further studies. Methods Based on the nuclear magnetic resonance image of one patient with mild stenosis of internal carotid artery, the lumped parameter model of the circle of Willis and the three-dimensional model of internal carotid artery were constructed. Those two different multi-scale models were constructed by unidirectional and bidirectional coupling. Results With the increase of stenosis degree, the inlet and outlet blood pressure and the outlet blood flow of internal carotid artery all decreased under two kinds of coupling method. The distribution of low time average wall shear stress (TAWSS) and high oscillatory shear index (OSI) of the internal carotid artery both increased with the increase of stenosis degree under two kinds of coupling method in general. The anterior cerebral artery segment showed lower shear stress and higher OSI with bidirectional coupling in 70% stenosis, and the blood flow direction of posterior communicating artery was changed, which was significantly different from unidirectional coupling results. Conclusions At a low degree of stenosis, the result of those two kinds of coupling method were consistent in general, but there was a significant difference in 70% stenosis, and the result of bidirectional coupling was closer to physiological parameters. The research findings can be better applied to the hemodynamic study of cerebrovascular diseases.