Objective To develop a highly sensitive and rapid nucleic acid detection method for the severe acute respiratory syndrome coronavirus 2(SARS-CoV-2). Methods We designed,developed,and manufactured an integrated disposable device for SARS-CoV-2 nucleic acid extraction and detection.The precision of the liquid transfer and temperature control was tested.A comparison between our device and a commercial kit for SARS-Cov-2 nucleic acid extraction was performed using real-time fluorescence reverse transcription polymerase chain reaction(RT-PCR).The entire process,from SARS-CoV-2 nucleic acid extraction to amplification,was evaluated. Results The precision of the syringe transfer volume was 19.2±1.9 μL(set value was 20),32.2±1.6(set value was 30),and 57.2±3.5(set value was 60).Temperature control in the amplification tube was measured at 60.0±0.0 ℃(set value was 60)and 95.1±0.2 ℃(set value was 95)respectively.SARS-Cov-2 nucleic acid extraction yield through the device was 7.10×106 copies/mL,while a commercial kit yielded 2.98×106 copies/mL.The mean time to complete the entire assay,from SARS-CoV-2 nucleic acid extraction to amplification detection,was 36 min and 45 s.The detection limit for SARS-CoV-2 nucleic acid was 250 copies/mL. Conclusion The integrated disposable devices may be used for SARS-CoV-2 Point-of-Care test(POCT).

Objective:To analyze the clinical, genetic characteristics and follow-up data of Chinese patients with hypophosphatasia (HPP).Methods:A retrospective analysis was conducted on six children with HPP admitted to the Department of Endocrinology, Genetics and Metabolism in Beijing Children′s Hospital from October 2010 to January 2019. Summarized the clinical and follow-up data of all six patients, as well as the pathogenic variants of five children.Results:The serum alkaline phosphatase levels of all six children (five males and one female) were significantly reduced (2-49 U/L). The 6 patients aged from 2 months to 6 years and 4 months, 4 infantile HPP, 1 childhood HIP and 1 odonto HPP. The four patients with infantile HPP presented with anorexia, slow weight gain and hypercalcemia, whereas the one patient with childhood HPP and the other patient with odonto HPP had tooth loss. The patient with childhood HPP also manifested with motor dysfunction. Genetic testing was conducted for five patients and 4 unrelated Chinese families and revealed 10 variations in ALPL gene, including 7 missense variation, 1 insertion variation, 1 frameshift variation, 1 deletion variation.Of which 3 were novel (p.Y28C, p.268, F>L, p.A176V).One of the infantile patients lost follow-up and the other three deceased. The clinical conditions were much improved with medical intervention for patients with childhood, orodonto HPP.Conclusions:While HPP patients with different ages of onset present with common features, the prognosis differ significantly. The prognosis is good for patients with childhood, orodonto HPP and poor for patients with infantile HPP. Genetic testing is the main method for definitive diagnosis.

Objective:To investigate the clinical and genetic features, and treatment of X-linked hypophosphatemic rickets (XLH).Methods:In this retrospective study, we reviewed the medical records of 25 pediatric patients with XLH who were admitted to Department of Endocrinology Genetics and Metabolism,Beijing Children′s Hospital from January 2010 to January 2020. The clinical characteristics, PHEX gene variants, as well as clinical outcome of the patients were summarized. To analyze the correlation between genotype and phenotype, the patients were divided into different subgroups according to the location of the variants, including N-terminal-located vs. C-terminal-located variant, and Zn-binding domain exon 17 or 19 variant vs. non-exon 17 or 19 variant. The age at onset, height standard deviation score (HtSDS), intercondylar or intermalleolar distance, fasting serum phosphorus, and HtSDS and intercondylar or intermalleolar distance at the final follow-up were compared by rank sum test or t text. Results:Among the 25 children with XLH, 8 were boys and 17 were girls. The median age of onset was 1.2 (1.0, 1.8) years, and the median age of diagnosis was 2.5 (1.5, 4.3) years. The main clinical manifestations were abnormal gait and lower limb deformity. The HtSDS was -2.0(-3.2, -0.8), and the intercondylar or intermalleolar distance was 4.5 (3.0, 6.0) cm. The fasting serum phosphorus level was 0.8 (0.7, 0.9) mmol/L, while the serum alkaline phosphatase level was (721±41) U/L and the serum calcium level was (2.5±0.1) mmol/L. Three patients (12%) had parathyroid hormone levels above the upper limit of the normal range. Twenty-five patients (100%) showed radiographic changes of active rickets. Nephrocalcinosis was found in 2 cases (9%). Twenty-four different PHEX variations were detected in 25 patients, among whom 11 (44%) had not been reported previously. No hot spot variation was found. No statistical differences (all P>0.05) were identified in clinical features and outcomes either in comparing patients with N-terminal (21 cases) and C-terminal (4 cases) variants, or in comparing patients with variant located in exon 17 or 19 (4 cases) or not (21 cases). Twenty-four cases (96%) were treated regularly with phosphate supplements and active vitamin D. After 2.7 (1.6, 5.0) years of follow-up, clinical symptoms were relieved in 96% (24/25) of the patients. The HtSDS after treatment had no significant difference compared to that before treatment (-2.0(-3.2, -0.8) vs.-2.0(-2.8, -1.1), Z =-0.156, P>0.05), while the intercondylar or intermalleolar distance after treatment was significantly reduced compared to that before treatment (4.5(3.0, 6.0) vs. 1.5(0, 3.3) cm, Z =-3.043, P<0.05). Bone X-rays were reexamined in 17 cases after treatment, and radiographic signs of rickets were improved. Eighteen cases had secondary hyperparathyroidism and 7 cases had nephrocalcinosis. Conclusions:The main clinical manifestations of XLH are abnormal gait, lower limb deformity and short stature. A high proportion of novel variations of PHEX gene but no hot spot variation neither genotype-phenotype correlation are found. Regular treatment with phosphate supplements and active vitamin D can significantly improve the symptoms except for the height. However, the rate of adverse events including secondary hyperparathyroidism and nephrocalcinosis seems to be high.

Objective To evaluate the factors related to contralateral hip fracture in the elderly patients with hip fracture by meta analysis.Methods Pubmed,Cochrane,CBMdisc,CNKI Chinese Journal Full Text Database and Wan Fang Database were searched for publications at home and abroad from January 2005 to April 2018 on factors related to contralateral hip fracture after hip fractures in the elderly.The publication quality was strictly evaluated before the data were extracted concerning gender and age(＞ 65years) of the patients,concomitant osteoporosis (Singh sign ≥ 4),primary fracture type,concomitant Parkinson disease,concomitant stroke,concomitant senile dementia,concomitant cataract,concomitant rheumatoid arthritis,concomitant diabetes,type of internal fixation for primary fracture and therapeutic compliance.Revman5.0 was used to perform the statistical analysis and the OR value and 95％ CI were calculated fore each index.Results A total of 17 studies were included involving 1,504 patients with contralateral hip fracture among 13,717 elderly patients with hip fracture.The factors related to the refracture of the contralateral hip were the age of the patients (OR ＝-3.55,95％ CI:-5.60 ～-1.50,P ＜ 0.001),osteoporosis (OR=2.38,95％CI:1.36～4.17,P=0.002),Parkinson disease (OR=4.54,95％CI:2.74～7.53,P ＜0.001),stroke (OR=0.33,95％ C I:0.18～0.59,P ＜ 0.001),senile dementia (OR=0.43,95％CI:0.29～0.62,P ＜0.001),cataract (OR=0.37,95％CI:0.22～0.63,P ＜0.001),rheumatoid arthritis (OR =0.32,95％ CI:0.21 ～ 0.50,P ＜ 0.001),diabetes (OR =0.65,95％ CI:0.47～0.91,P=0.01),type of internal fixation for primary fracture (OR=0.51,95％ CI:0.30 ～ 0.85,P =0.01),and therapeutic compliance (OR =0.36,95％ CI:0.21 ～ 0.64,P ＜ 0.001).However,the refracture of the contralateral hip was not related to gender (OR =1.07,95％ CI:0.45 ～2.56,P=0.88),smoking (OR=0.86,95％CI:0.40～1.86,P=0.70),fracture type (OR=0.97,95％ CI:0.60～1.57,P=0.90),or hypertension (OR=0.70,95％ C I:0.41～1.21,P=0.20).Conclusions In elderly patients with hip fracture,the risks for contralateral hip fracture may be advanced age,concomitant osteoporosis,Parkinson disease,stroke,senile dementia,cataract,rheumatoid arthritis and diabetes,type of internal fixation for primary fracture,and poor therapeutic compliance.However,no sufficient evidence has suggested that gender,smoking,type of hip fracture or concomitant hypertension might be associated with the contralateral hip fracture.

Objective:To assess the long-term effectiveness and safety of rituximab (RTX) for the treatment of pemphigus, and to evaluate the effect of RTX on immune indices.Methods:A retrospective study was conducted, and patients with pemphigus who received monotherapy or combination therapy with RTX (375 mg/m 2 body surface area, once a week for 4 consecutive weeks) were collected from the Department of Dermatology, Peking University First Hospital from February 2008 to July 2017. Levels of autoantibodies and proportion of B cells in patients were determined at baseline and different follow-up time points, and their changes and relationship with therapeutic effect were analyzed. Time-to-event outcomes (disease control, complete remission and relapse) were estimated using the Kaplan-Meier method. The median ( M) as well as 25th ( P25) and 75th ( P75) percentile values were calculated for repeatedly measured immune indices (autoantibodies and B cells) , and the median level of immune indice-time curve was drawn. Results:A total of 53 patients of Han nationality with pemphigus were included, including 40 with pemphigus vulgaris and 13 with pemphigus foliaceus. The male to female ratio was 0.96∶1, the median age was 37.4 years, and the median duration of disease was 13.4 months at baseline. The median follow-up duration ( P25, P75) was 37.5 (25.0, 54.7) months. Forty-eight (90.6%) patients achieved disease control, and the time to disease control was 1.7 (1.1, 3.2) months. Thirty-eight (71.7%) patients achieved complete remission, and the time to complete remission was 13.1 (9.6, 27.5) months. During the follow up, 12 of the 38 (31.6%) patients who had complete remission experienced recurrence, with the time to recurrence being 12.4 (4.8, 19.8) months. The median immune indice level-time curve showed that anti-Dsg1 and Dsg3 autoantibody levels decreased when skin lesions resolved, but increased when skin lesions relapsed. The most common severe adverse reaction was pulmonary infection, with a mortality rate of 3.8% (2/53) . Conclusions:RTX shows marked long-term effectiveness for the treatment of pemphigus. Pulmonary infection during treatment is worthy of the highest attention. The autoantibody levels can serve as an index for evaluating the effectiveness of RTX in the treatment of pemphigus.

Objective: To investigate the clinical effect of combined anterior and posterior approach revision on complex acetabular fractures. Methods: A retrospective case series study was performed on the clinical data of 21 patients with complex acetabular fractures who underwent failed surgery through single approach from June 2012 to June 2017. There were 16 males and five females, averagely aged 34.6 years (range, 24-45 years). According to Letournel-Judet classification, there were seven patients with transverse+ posterior wall fracture, five patients with anterior column+ posterior semi-transverse fracture, four patients with double column fracture and five patients with "T" fracture. The first operation was performed by ilioinguinal approach in nine patients and by Kocher-Langebeck (K-L) approach in 12 patients. Revision surgery was performed using a combined anterior and posterior approach. The operation time and intraoperative blood loss were recorded, and the fracture healing was observed. The quality of fracture reduction was assessed according to Matta reduction criteria, and hip function by the modified D Aubigne and Postel score. The complications during and after operation were recorded. Heterotopic ossification was evaluated according to Brooker's criteria. Results: The patients were followed up for 12 to 36 months, with an average of 27 months. The operation time ranged from 180 to 360 minutes, with an average of 270 minutes. Intraoperative bleeding was 1 000-3 800 ml, with an average of 2 000 ml. Fractures were healed, with the healing time ranging from 3.5 to 7 months, with an average of 5 months. According to Matta reduction criteria, there were eight patients with anatomical reduction, 12 with satisfactory reduction and one with unsatisfactory reduction. The improved D Aubigne and Postal score was (11.1±1.9)points preoperatively and (15.6±1.7)points six months after operation (P<0.05), the outcome of hip function was excellent in three patients, good in 14, fair in three, and poor in one, with the excellent and good rate of 81%. There were three patients with transient injury of sciatic nerve, one patient with traumatic arthritis and one with heterotopic ossification of Brooker II. No femoral head necrosis or deep venous thrombosis in the lower extremities was found. Conclusion For complex acetabular fractures, combined anterior and posterior approach revision can promote fracture reduction, fracture healing, and functional recovery, with low incidence of complications.

N-methyladenosine (mA), catalyzed by the methyltransferase complex consisting of Mettl3 and Mettl14, is the most abundant RNA modification in mRNAs and participates in diverse biological processes. However, the roles and precise mechanisms of mA modification in regulating neuronal development and adult neurogenesis remain unclear. Here, we examined the function of Mettl3, the key component of the complex, in neuronal development and adult neurogenesis of mice. We found that the depletion of Mettl3 significantly reduced mA levels in adult neural stem cells (aNSCs) and inhibited the proliferation of aNSCs. Mettl3 depletion not only inhibited neuronal development and skewed the differentiation of aNSCs more toward glial lineage, but also affected the morphological maturation of newborn neurons in the adult brain. mA immunoprecipitation combined with deep sequencing (MeRIP-seq) revealed that mA was predominantly enriched in transcripts related to neurogenesis and neuronal development. Mechanistically, mA was present on the transcripts of histone methyltransferase Ezh2, and its reduction upon Mettl3 knockdown decreased both Ezh2 protein expression and consequent H3K27me3 levels. The defects of neurogenesis and neuronal development induced by Mettl3 depletion could be rescued by Ezh2 overexpression. Collectively, our results uncover a crosstalk between RNA and histone modifications and indicate that Mettl3-mediated mA modification plays an important role in regulating neurogenesis and neuronal development through modulating Ezh2.
Adenosine ; analogs & derivatives ; metabolism ; Adult Stem Cells ; cytology ; metabolism ; Animals ; Brain ; metabolism ; Cell Differentiation ; genetics ; Cell Proliferation ; Enhancer of Zeste Homolog 2 Protein ; metabolism ; Gene Expression Regulation ; Methyltransferases ; metabolism ; Mice, Inbred C57BL ; Neural Stem Cells ; cytology ; metabolism ; Neurogenesis ; genetics ; Neurons ; cytology ; metabolism ; RNA, Messenger ; genetics ; metabolism

A 2-month-and-20-day-old girl was hospitalized because of slow weight gain for 20 days.The patient was diagnosed as anemia,pneumonia and neutrophils firstly,and showed no improvement during anti-infective therapy,blood transfusion,and leukopenia drugs.The urinary organic acid spectrum of the patient showed increased methylmalonic acid slightly,but the normal value was found by 4 tests,which was inconsistent with the typical methylmalonic aciduria.By analyzing clinical manifestations and plasma homocysteine,folic acid and vitamin B12 levels,she was considered to have intracellular cobalamin metabolic disorder.Then,normal diet,intramuscular injection of vitamin B12,and feeding calcium,betaine,and L-carnitine were given,and the girl's symptoms were improved significantly.Latter gene analysis further showed that the patient had methylmalonic aciduria CblC type.Clinical manifestations of methylmalonic aciduria are complex and individually various,different therapies showed be applied to different causes.The value of urinary methylmalonic acid level alone cannot judge etiology;diagnosis should be based on blood amino acids and carnitine spectrum,homocysteine level and vitamin B12 level.

Objective: To study the effect of different dissolution media on the dissolution curve of risperidone oral soluble films to provide reference for the quality evaluation of the preparation.

Objective To modified doxorubicin liposome with transferrin(TF),and to investigate its inhibition efficacy on the proliferation of human breast cancer cells.Methods The liposome was prepared by thin film ultrasonic,and doxorubicin liposomal was prepared by sulfuric acid gradient.The TF-doxorubicin lipo-some was prepared by the post insertion method.The uptake of TF-liposomal doxorubicin on breast cancer cells MCF-7 and MDA-MB-231 were detected by confocal microscopy.The killing ability of TF-doxorubicin liposomal targeting for MCF-7 and MDA-MB-231 were detected by MTT assay.Inhibitory effect of TF-doxorubicin lipo-some on the growth of MCF-7 and MDA-MB-231 were detected by soft agar colony assay.Results Confocal microscopy result showed that the uptake of TF-liposomal doxorubicin on MCF-7 and MDA-MB-231 were signifi-cantly higher than doxorubicin liposomal.Cell-killing ability on MCF-7 and MDA-MB-231 showed that the IC50 in TF-liposomal doxorubicin [MCF-7 cells:(20.8 ±3.2)μmol/L;MDA-MB-231 cells:(20.1 ±3.0)μmol/L)] were significantly lower than the liposomal [(1 58.6 ±24.6)μmol/L;(1 60.1 ±25.1 )μmol/L)]and free doxorubicin [(1 61 .7 ±26.2)μmol/L;(1 66.9 ±27.0)μmol/L)],with significant differences(F =1 1 6.03, P <0.001 ;F =75.29,P <0.001 ).Soft agar colony assay showed that the inhibition of TF-doxorubicin lipo-some on colony growth were significantly higher than doxorubicin liposome,free doxorubicin and control [dia-meter of MDA-MB-231 cells:(60.5 ±10.4)μm,(94.3 ±16.8)μm,(1 31 .8 ±22.6)μm,(162.8 ±30.3)μm;diameter of MCF-7 cells:(31 .8 ±5.5)μm,(62.1 ±11 .1 )μm,(108.6 ±1 8.6)μm,157.4 ±29.3)μm],with significant differences (F =87.17,P <0.000 1 ;F =178.23,P <0.000 1 ).Conclusion TF-doxorubicin lipo-some has a significant inhibitory effect on the proliferation of breast cancer cells in vitro,and can effectively and specifically kill the breast cancer cells,which provides theoretical basis for the treatment of breast cancer in vivo.