Tropane alkaloids (TAs) are a class of anticholinergic drugs widely used in clinical practice and mainly extracted from plant, among which Atopa belladonna is the main commercial drug source. It is of great industrial value to obtain TAs in large quantities by plant metabolic engineering. In TAs pathway, cytochrome oxidase CYP82M3 catalyze the synthesis of tropinone and then tropinone reductase I (TRI) compete with TRII for tropinone to form tropine leading to the TAs synthesis (drainage). In this study, based on the "increasing flow and drainage" metabolic engineering strategy, two genes, namely HnCYP82M3 and DsTRI from Hyoscyamus niger and Datura stramonium, respectively, were overexpressed in the hair roots of A. belladonna, with a view to promote the TAs accumulation. The HnCYP82M3 gene was cloned from the root of H. niger, and it encoded amino acid with 91.7% sequence identity with AbCYP82M3 from A. belladonna. Overexpression of HnCYP82M3 alone did not affect the content of TAs in hair roots of A. belladonna, indicating that CYP82M3 was not a key enzyme in TAs biosynthesis. Simultaneous overexpression of HnCYP82M3 and DsTRI greatly promoted the accumulation of the three TAs, and the contents of hyoscyamine, anisodamine and scopolamine were 4.97 times, 2.83 times and 2.19 times that of the control, respectively, and the increase amplitude was greater than that of single overexpression of DsTRI. This study showed that the "increasing flow and drainage" strategy of enzyme genes co-expression at branch points was a promising metabolic engineering method to effectively improve the biosynthesis of TAs in A. belladonna, and laid a theoretical and technical foundation for the large-scale industrial acquisition of TAs.

The rol genes on pRiA4 plasmid of Agrobacterium rhizogenes are potent genes that promote secondary metabolism. Molecular breeding of Atropa belladonna can be conducted by introducing rol genes to increase tropane alkaloids (TAs) content in A. belladonna. In this study, the rolB gene was overexpressed in A. belladonna plants to study the effect of rolB gene on the biosynthesis of TAs. The phenotype, TAs content and expression levels of key enzyme genes in the pathway of TAs biosynthesis of transgenic A. belladonna were analyzed. The results showed that transgenic A. belladonna had developed root system, enlarged leaves, increased leaf fresh weight, deepened leaf color, enlarged flowers, changed flower shape, reduced pistil height and decreased pollen vitality. The content of TAs in the stems of transgenic A. belladonna was significantly higher than that of the control, and the contents of scopolamine, anisodamine, hyoscyamine can reach 2.11-2.91, 1.23-2.37 and 4.88-5.20 times of the control, respectively. Compared with the control group, the expressions of key enzymes putrescine N-methyltransferase (PMT), type III polyketide synthase (PYKS), tropinone reductase I (TRI), aromatic amino acid aminotransferase 4 (ArAT4), UDP-glycosyltransferase 1 (UGT1) and hyoscyamine 6-β-hydroxylase (H6H) in the TAs biosynthesis pathway were up-regulated, and the expression of tropinone reductase II (TRII) as a metabolic shunting gene was down-regulated. The results indicated that rolB gene enhanced TAs synthesis ability in roots and accumulation in stems of A. belladonna by enhancing metabolic flow of TAs synthesis pathway and weakening the metabolic shunt of competing pathway. This study laid a foundation for molecular breeding of A. belladonna with high-yield TAs content using rolB gene.

The paper explores the evolution of "bone-approaching" acupuncture, its effect target and mechanism. The concrete operation procedure of "bone-approaching" method is recorded originally in Huangdi Neijing (Inner Canon of Yellow Emperor) as short needling and Shu needling (referring to the category of the five needling technique). The periosteum is the most effective stimulation target of "bone-approaching" acupuncture for analgesia, regaining consciousness and regulating spirit. The "bone-approaching" acupuncture is not only prominently effective on bone bi syndrome, but also has the unique effect on painful, encephalogenic and emotional diseases. The paper summarizes and improves "bone-approaching" acupuncture, i.e. "touching bone surface" with needle tip by slow insertion, "touching bone surface" without pain by swift insertion and "touching bone" with needle body by oblique insertion. It contributes to the inheritance, development and supplementation to the bone needling techniques in Huangdi Neijing and is significant for broadening the clinical application range of acupuncture.
Humans ; Acupuncture Therapy ; Periosteum ; Analgesia ; Pain Management ; Consciousness ; Pain

To analyze the treatment and prognosis of T cell acute lymphoblastic leukemia(T-ALL)in adults. Method The clinicobiogical and survival data of 68 adult patients with newly diagnosis T-ALL were retrospectively analzyed. Results The median age of these 68 patients was 23 years(14-60 years).T-ALL was more common in men(81%).After the first cycle of treatment,complete remission was achieved in 50 patients(73%).The highest complete remission(CR) rate was in patients with cortex T-ALL(100%),followed by other T-ALL(73%)and early T-cell precursor lymphoblastic leukemia(54%),(=5.712,=0.058).The CR rate for adults aged >35 years was significantly lower than that of patients aged ≤ 35 years(40% 79%,=6.364,=0.012).The overall CR rate after the second treatment course was 93%.For patients treated with chemotherapy,autograft hematopoietic stem cell transplantation(auto-SCT),and allogeneic SCT,the median relapse free survival was 10 months,24 months,and not reached,respectively(=0.002).The 5-year overall survival rate was 25% for all patients;for patients treated with chemotherapy,auto-SCT and allogeneic SCT,the median overall survival was 24 months,34 months,and 30 months,respectively(=0.007),and the 5-year overall survival rate was 9%,33%,and 38%(=0.037).Multivariate analysis showed leukocyte count ≥100×10 /L was a risk factor for decreased relapse free survival(risk ratio 2.540,95%=1.058-6.099,=0.037). Conclusion Adult T-ALL patients have poor prognosis,which may be improved by SCT.
Adolescent ; Adult ; Disease-Free Survival ; Female ; Hematopoietic Stem Cell Transplantation ; Humans ; Male ; Middle Aged ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; diagnosis ; therapy ; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma ; diagnosis ; therapy ; Prognosis ; Remission Induction ; Retrospective Studies ; Survival Rate ; Treatment Outcome ; Young Adult

Objective: To investigate the prognostic significance of detection of minimal residual disease after first induction treatment (MRD(1)) in adult acute lymphoblastic leukemia (ALL) patients treated with autologous stem cell transplantation (auto-HSCT). Methods: The clinical data of 87 ALL patients who underwent auto-HSCT during February 2006 to April 2017 with MRD(1) detection data by flow cytometry were analyzed retrospectively. The relationship between MRD(1) and relapse and survival of ALL patients after auto-HSCT was studied. Results: Of 87 patients, 26 (29.9%) were MRD(1) positive. The proportion of high-risk immunophenotype (pro-B, pro-T, pre-T, mature T) was significantly higher in MRD(1)-positive patients than that in MRD(1) negative patients (34.6% vs 14.5%, P=0.038). There was no significant difference between positive and negative MRD(1) patients at age, sex, lineage (T/B), immunophenotype (standard risk/high risk), high white blood cell count (B-ALL>30×10(9)/L or T-ALL>100×10(9)/L), high-risk chromosome/gene ratio, the time from first complete remission to transplantation and pre-treatment regimen. The 5-year overall survival (OS) and leukemia-free survival (LFS) in MRD(1) negative and positive patients were 72.7% vs 47.3% (P=0.004) and 75.7% vs 29.6% (P<0.001), respectively. Multivariate analysis showed that positive MRD(1) was an independent risk factor for OS (HR=3.007, 95% CI 1.256-7.200, P=0.013) , and positive MRD(1) and high-risk immunophenotype were risk factors for LFS (HR=3.986, 95% CI 1.813-8.764, P=0.001; HR=2.981, 95% CI 1.373-6.473, P=0.006) . Conclusions: Auto-HSCT could not reverse the poor prognosis of MRD(1) positive patients. Auto-HSCT treatment is optional for patients with MRD(1) negative and maintaining MRD(1) negative status during intensive therapy.
Adult ; Hematopoietic Stem Cell Transplantation ; Humans ; Neoplasm, Residual ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; Prognosis ; Retrospective Studies ; Transplantation, Autologous

BACKGROUNDHalo nevus (HN) has been shown to be associated with vitiligo, but no standard signs are currently available to identify HN patients at risk of vitiligo, and the relevant data obtained in previous studies are somewhat conflicting. This study aimed to identify factors affecting the presence of vitiligo in HN patients.

METHODSWe performed a retrospective study on consecutive patients with HN at the First Affiliated Hospital of Sun Yat-sen University between January 2011 and December 2016. Detailed demographic and clinical data were collected to identify the factors associated with the presence of vitiligo in this cohort of patients using uni- and multi-variate logistic regression analyses.

RESULTSA total of 212 HN patients were included, 101 of whom had vitiligo-associated HN (HNV). Univariate analysis indicated that a personal history of thyroid diseases was positively associated with HNV (odds ratio [OR] = 10.761, P = 0.025), while the onset age of HN was negatively associated with HNV (OR = 0.537, P = 0.026). Multivariate analysis demonstrated that the Koebner phenomenon (KP; OR = 10.632, P < 0.0001), multiple HN (OR = 3.918, P < 0.0001), and a familial history of vitiligo (OR = 3.222, P = 0.014) were independent factors associated with HNV.

CONCLUSIONSHN without vitiligo has clinical features distinct from HN associated with vitiligo. HN patients with KP, multiple lesions, or familial history of vitiligo are more likely to develop vitiligo and therefore should be monitored for clinical signs of such accompanied conditions.

OBJECTIVETo study the effect of β8 expression on transforming growth factor β1(TGF-β1) activation in astrocytes with oxygen glucose deprivation (OGD).

METHODSAstrocytes were cultured and then subjected to OGD to generate hypoxia-ischemia (HI) model in vitro. Immunocytochemistry was used to detect the expression and distribution of β8 in nomoxia cultured cells. β8 protein expression was quantified by Western blot at 12 hours, 1 day and 2 days after OGD. Astrocytes and luciferase reporter cells (TMLC) were co-cultured. β8 RNA interference system was established to specifically inhibit β8 expression in cultured astrocytes. TGF-β1 activation was then detected in the co-culture system.

RESULTSβ8 was mainly located in the cytoplasm and neurites of astrocytes. OGD resulted in increase of β8 protein expression at 12 hours after reoxygenation in astrocytes, which was peaked at 1 day after reoxygenation. TGF-β1 activation was in accordance with β8 expression in astrocyte-TMLC co-culture system after reoxygenation. After the inhibition of β8, TGF-β1 activation was significantly reduced in all time points.

CONCLUSIONSThe highly expressed β8 plays important roles in the regulation of TGF-β1 activation in neonatal rats with hypoxic-ischemic brain damage.

Animals ; Astrocytes ; metabolism ; Female ; Glucose ; metabolism ; Hypoxia-Ischemia, Brain ; metabolism ; Integrin beta Chains ; physiology ; Male ; Oxygen ; metabolism ; Rats ; Rats, Sprague-Dawley ; Transforming Growth Factor beta1 ; metabolism

Inhaled NO (iNO) has been shown to have beneficial effects on decreasing pulmonary inflammation, increasing function of surfactant and improving lung growth in prematurely born animal models. iNO has been gradually applied in the neonatal intensive care unit since its first use for persistent pulmonary hypertension (PPHN) in the early 1990's. Although many research findings have shown the benefits of iNO for hypoxic respiratory failure (HRF) of preterm infants, there is no certain evidence to support the routine use of iNO in premature infants. According to recent literature, the mechanism of iNO therapy, treatment scheme, iNO effectiveness and safety in premature infants were reviewed in this article, so as to provide bases for the clinical use of this treatment.
Administration, Inhalation ; Humans ; Hypoxia ; complications ; Infant, Newborn ; Infant, Premature ; Nitric Oxide ; administration & dosage ; adverse effects ; Respiratory Insufficiency ; drug therapy

OBJECTIVETo explore the role and mechanisms of FOXO3a nuclear translocation in neuronal apoptosis after hypoxia-ischemia (HI).

METHODSOne hundred and sixty 10-day-old Sprague-Dawly rats were randomly divided into two groups: HI and sham-operated. The right common carotid artery was ligated followed by hypoxia exposure for 2.5 hours in the HI group. The sham-operated group rats were not subjected to carotid artery ligation or hypoxia treatment. Rat cerebral cortex was collected at 0.5, 2, 4, 8 and 24 hours after hypoxia. Western blot was used to detect expression of total FOXO3a protein, pnuclear and cytoplasmic FOXO3a and Bim. TUNEL staining was used to detect apoptotic cells.

RESULTSThe nuclear protein of FOXO3a obviously increased from 0.5 to 24 hours after HI in a time-dependent manner compared with the sham-operated group (P<0.01). On the contrary, cytoplasmic protein evidently decreased from 0.5 to 24 hours in the HI group compared with the sham-operated group (P<0.01). Bim protein increased from 0.5 hour, peaked at 2 hours, started to decline at 4 hours (P<0.01), and returned to baseline level at 8 and 24 hours after HI in the HI group compared with the sham-operated group. TUNEL positive cells started to express at 4 hours, and peaked at 24 hours after HI (P<0.01). However, TUNEL positive cells were rarely found in the sham-operated group.

CONCLUSIONSHI induces FOXO3a translocation from cytoplasm to nucleus, and enhances protein expression of its target gene Bim in the neonatal rat brain. The upregulation of Bim expression might be related to neuronal apoptosis.

Animals ; Animals, Newborn ; Apoptosis ; Apoptosis Regulatory Proteins ; analysis ; Bcl-2-Like Protein 11 ; Female ; Forkhead Box Protein O3 ; Forkhead Transcription Factors ; genetics ; physiology ; Hypoxia-Ischemia, Brain ; pathology ; Male ; Membrane Proteins ; analysis ; Neurons ; pathology ; Proto-Oncogene Proteins ; analysis ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo analyze the measles immunity level of persistent population in Beijing.

METHODSA total of 2125 objects from 10 age groups, who had been living in Beijing for over 6 months, were selected from urban and rural areas in Beijing in 2012. Demographic characteristics, history of measles and vaccine immunization were investigated by questionnaire. 5 ml blood sample of each subject was collected, and the Measles IgG antibody was measured by ELISA assay.

RESULTSPositive rate of measles antibody was 84.71% (1800/2125) and standardized positive rate was 88.07% . Median of antibody was 960.46 IU/L. Positive rate and median of measles antibody were significantly different between population from different age groups (χ(2) = 341.60, P < 0.01; H = 216.27, P < 0.01). Antibody positive rate and median were lowest in the <1 year age group, which were separately 43.06% (90/209) and 185.80 IU/L; and highest in the 1-4 (97.31% (181/186) and 2448.81 IU/L) and 5-9 years age group (96.46% (218/226) and 1910.72 IU/L). The range of antibody positive rate and median in adults of ≥ 15 years were 81.98%-90.14% and 744.38-1474.84 IU/L. Antibody positive rate and median in persistent population, which were separately 82.45% (883/1071) and 899.82 IU/L, were lower than those in migrant population, which were 87.00% (917/1054) and 166.19 IU/L, respectively (χ(2) = 8.51, P < 0.01;U = 538 704.00, P < 0.01). Antibody positive rate and median in population with vaccination history, which were separately 91.95% (891/969) and 1443.11 IU/L, were higher than those population without vaccination history and people whose history unknown (32.95% (57/173) , 127.33 IU/L; 86.67% (852/983) , 923.73 IU/L). The difference showed statistical significance (χ(2) = 399.92, P < 0.01; H = 202.11, P < 0.01).

CONCLUSIONAmong the persistent population in China, measles antibody level among the children aging 1-9 years old was high enough to prevent outbreak and epidemic of measles. However, we should try our best to strengthen the measles antibody level among the babies younger than 1 year old and the migrant population aging between 15 and 40 years old.

Adolescent ; Adult ; Antibodies, Viral ; blood ; Child ; Child, Preschool ; China ; epidemiology ; Female ; Humans ; Infant ; Male ; Measles ; epidemiology ; immunology ; prevention & control ; Measles virus ; Young Adult