Exosomes are a subpopulation of extracellular vesicles ranging in size from 30 to 150 nm. They contain a variety of biological molecules including proteins, lipids, and genetic materials by which they can act as mediators of cell-to-cell communication. Increasing studies on exosomes have elucidate the mechanism of communication between immune cells. Mast cells are found to release exosomes in both resting and activated states, but the quantity, contents and functions of the exosomes are significantly different in different states. Mast cell-derived exosomes are involved in the maturation and antigen presentation of dendritic cells, and mediate the activation of T lymphocytes and the polarization of macrophages. These results provide new insights into the role of exosomes in mast cell function and immune regulation.

Objective:To detective the cut-off values of amino acid levels in premature infants in Sichuan.Methods:Data of newborns screening for inherited metabolic diseases (IMD) by tandem mass spectrometry in Sichuan Province from January 2018 to December 2019 were retrospectively analyzed.They were divided into premature infant group ( n=2 264, 1 312 males and 952 females) and full-term infant group ( n=53 275, 28 269 males and 25 006 females). The cut-off values of amino acids in dry blood spots were expressed as percentage ( P0.5 - P99.5), and rank sum test was used for comparison between preterm and full-term infants. Results:(1) The distribution of 11 amino acids [alanine (ALA), arginine (ARG), citrulline (CIT), glycine(GLY), leucine (LEU), methionine (MET), ornithine (ORN), phenylalanine (PHE), proline (PRO), tyrosine (TYR) and valine (VAL)] in premature infants were abnormal.(2) The cut-off values of amino acids in premature infants were as follows: ALA: 135.20-552.33 μmol/L, ARG: 1.34-47.04 μmol/L, CIT: 5.66-32.02 μmol/L, GLY: 181.48-909.93 μmol/L, LEU : 71.10-283.29 μmol/L, MET: 4.21-34.51 μmol/L, ORN: 40.58-293.76 μmol/L, PHE: 23.60-106.30 μmol/L, PRO: 77.76-358.24 μmol/L, TYR: 27.52-352.91 μmol/L, VAL: 53.74-228.37 μmol/L.(3) The cut-off values of amino acid in full-term infants were as follows: ALA: 135.20-552.33 μmol/L, ARG: 1.30-42.73 μmol/L, CIT: 5.92-30.35 μmol/L, GLY: 208.17-980.09 μmol/L, LEU: 72.91-287.49 μmol/L, MET: 4.27-33.90 μmol/L, ORN: 48.40-305.59 μmol/L, PHE: 27.63-92.27 μmol/L, PRO: 97.38-372.75 μmol/L, TYR: 40.19-276.54 μmol/L, VAL: 65.75-237.92 μmol/L.(4) Except for PHE ( Z=-0.58, P>0.05), the other indicators were significantly different between 2 groups [ALA ( Z=-15.32, P<0.05), ARG ( Z=-5.62, P<0.05), CIT ( Z=-5.86, P<0.05), GLY ( Z=-14.52, P<0.05), LEU ( Z=-5.62, P<0.05), MET ( Z=-5.22, P<0.05), ORN ( Z=-13.01, P<0.05), PRO ( Z=-22.09, P<0.05), TRY ( Z=-2.09, P<0.05), VAL ( Z=-17.82, P<0.05)]. Conclusions:The establishment of the cut-off values of amino acids in premature infants in Sichuan provides a theoretical basis for laboratory diagnosis of IMD screening, which enhances the accuracy of diagnosis and avoids excessive medical treatment.

Objective:To establish a disease risk prediction model for the newborn screening system of inherited metabolic diseases by artificial intelligence technology.Methods:This was a retrospectively study. Newborn screening data ( n=5 907 547) from February 2010 to May 2019 from 31 hospitals in China and verified data ( n=3 028) from 34 hospitals of the same period were collected to establish the artificial intelligence model for the prediction of inherited metabolic diseases in neonates. The validity of the artificial intelligence disease risk prediction model was verified by 360 814 newborns ' screening data from January 2018 to September 2018 through a single-blind experiment. The effectiveness of the artificial intelligence disease risk prediction model was verified by comparing the detection rate of clinically confirmed cases, the positive rate of initial screening and the positive predictive value between the clinicians and the artificial intelligence prediction model of inherited metabolic diseases. Results:A total of 3 665 697 newborns ' screening data were collected including 3 019 cases ' positive data to establish the 16 artificial intelligence models for 32 inherited metabolic diseases. The single-blind experiment ( n=360 814) showed that 45 clinically diagnosed infants were detected by both artificial intelligence model and clinicians. A total of 2 684 cases were positive in tandem mass spectrometry screening and 1 694 cases were with high risk in artificial intelligence prediction model of inherited metabolic diseases, with the positive rates of tandem 0.74% (2 684/360 814)and 0.46% (1 694/360 814), respectively. Compared to clinicians, the positive rate of newborns was reduced by 36.89% (990/2 684) after the application of the artificial intelligence model, and the positive predictive values of clinicians and artificial intelligence prediction model of inherited metabolic diseases were 1.68% (45/2 684) and 2.66% (45/1 694) respectively. Conclusion:An accurate, fast, and the lower false positive rate auxiliary diagnosis system for neonatal inherited metabolic diseases by artificial intelligence technology has been established, which may have an important clinical value.

Objective:To explore the change characteristics of amino acid levels in neonates, so as to provide theoretical basis for accurate clinical interpretation.Methods:By preliminary screening and diagnosis from 32 855 newborns, 32 843 samples were collected using tandem mass spectrometry to inherited metabolic disease (IMD) scree-ning in Sichuan Province from January to December 2018.Afterwards, according to gestational age (1 363 premature infants, 31 468 full-term infants and 12 overdue infants), blood collection time (3-7 days old, 3 095 cases, 8-28 days old, 1 637 cases, and more than 28 days old, 248 cases) and season (7 737 cases in the first quarter, 11 428 cases in the second quarter, 5 482 cases in the third quarter, and 8 196 cases in the fourth quarter), neonates were divided into different study groups.The difference of amino acid level in each group was compared, and the correlation between various influencing factors and metabolic index was analyzed.Results:(1) The distribution of 11 amino acids [alanine(ALA), arginine(ARG), citrulline(CIT), glycine(GLY), leucine+ isoleucine+ hydroxyproline (LEU+ ILE+ PRO-OH), methionine(MET), ornithine(ORN), phenylalanine(PHE), proline(PRO), tyrosine(TYR), and valine(VAL)] in neonates showed non-normally distribution.(2)The distribution of 11 amino acids in different gestational age were tested by nonparametric test, except for PHE( H=0.61, P>0.05)and TYR( H=2.02, P>0.05), and other indicators were significantly different [ALA( H=187.11, P<0.05), ARG( H=23.60, P<0.05), CIT( H=22.90, P<0.05), GLY( H=85.18, P<0.05), LEU( H=56.42, P<0.05), MET( H=18.74, P<0.05), ORN( H=129.27, P<0.05), PRO( H=344.40, P<0.05), and VAL( H=272.92, P<0.05)]. (3) The distribution of 11 amino acids in different blood collection time were significantly different [ALA( H=65.19, P<0.05), ARG( H=404.48, P<0.05), CIT( H=502.13, P<0.05), GLY( H=1 719.44, P<0.05), LEU( H=396.41, P<0.05), MET( H=199.39, P<0.05), ORN( H=31.26, P<0.05), PHE( H=325.49, P<0.05), PRO( H=70.09, P<0.05), TYR( H=159.29, P<0.05), and VAL( H=102.52, P<0.05)]. (4) The distribution of 11 amino acids in different birth seasons were significantly different [ALA( H=401.37, P<0.05), ARG( H=3 229.03, P<0.05), CIT( H=65.45, P<0.05), GLY( H=597.82, P<0.05), LEU( H=1 120.42, P<0.05), MET( H=10 515.18, P<0.05), ORN( H=1 275.23, P<0.05), PHE( H=2 260.17, P<0.05), PRO( H=319.57, P<0.05), TYR( H=884.37, P<0.05), and VAL( H=1 824.49, P<0.05)]. Conclusion:According to different gestational age, season and blood collection time, the metabolism of amino acids in neonates was different.When using tandem mass spectrometry for detection, appropriate interpretation criteria should be selected based on different conditions.

OBJECTIVE: To investigate the mechanism by which epigallocatechin gallate (EGCG) induces gene demethylation and promotes the apoptosis of acute myeloid leukemia KG-1 and THP-1 cell lines. METHODS: KG-1 and THP-1 cells treated with 25, 50, 75, 100 or 150 μg/mL EGCG for 48 h were examined for gene methylation using MSP and for cell proliferation using MTT assay. The changes in cell cycle and apoptosis of the two cell lines after treatment with EGCG for 48 h were detected using flow cytometry. The mRNA and protein expressions of DNMT1, CHD5, p19, p53 and p21 in the cells were detected using RT-quantitative PCR and Western blot. RESULTS: EGCG dose-dependently reversed hypermethylation of gene and reduced the cell viability in both KG-1 and THP-1 cells ( < 0.05). EGCG treatment caused obvious cell cycle arrest in G1 phase, significantly increased cell apoptosis, downregulated the expression of DNMT1 and upregulated the expressions of CHD5, p19, p53 and p21 in KG-1 and THP-1 cells ( < 0.05). CONCLUSIONS EGCG reduces hypermethylation of gene in KG-1 and THP-1 cells by downregulating DNMT1 to restore its expression, which results in upregulated expressions of p19, p53 and p21 and induces cell apoptosis.

Objective To report retrograde transposition of composite tissue flap pedicled with peroneal artery in the treatment of bone and soft tissue defects following calcaneal osteomyelitis.Methods During the period from January 2008 through January 2015,47 patients with bone and soft tissue defects following calcaneal osteomyelitis were treated by retrograde transposition of composite tissue flap pedicled with peroneal artery.They were 43 men and 4 women,aged from 24 to 67 years(average,43.6 years).After radical focus debridement,the areas of skin defect ranged from 2 cm × 2 cm to 5 cm× 5 cm and the bone defects ranged from 1.5 cm × 1.0 cm × 2.0 cm to 3.0 cm × 3.0 cm × 4.5 cm.Results All the 47 patients were followed up for 6 to 72 months (average,25.2 months).Relapse of osteomyelitis occurred in 2 cases but secondary bone graft was needed in none.All the patients recovered their walking function.The texture and color of the composite tissue flaps were similar to those of normal skin,without obvious swelling.The flaps appeared good and imposed no obstacle to wearing shoes.The ankles and hind feet of the patients scored from 58 to 91 points (average,76.6 points) according to the American Orthopaedic Foot and Ankle Society (AOFAS) evaluation system,giving 7 excellent,32 good and 8 fair cases.Hallux flexion contracture without skin ulcer was observed in 10 cases 6 months after surgery,necessitating no surgical intervention.Conclusion Retrograde transposition of composite tissue flap pedicled with peroneal artery can effectively treat bone and soft tissue defects following calcaneal osteomyelitis,leading to a low recurrence rate and good foot appearance.

AIM:To study whether hydrogen sulfide (H2S) protects H9c2 cardiomyocytes against high glucose ( HG)-induced injury by inhibiting necroptosis .METHODS:The protein levels of RIP3 ( an indicator of necroptosis ) and cleaved caspase-3 were determined by Western blot .The cell viability was measured by CCK-8 assay.The intracellular le-vels of reactive oxygen species (ROS) were detected by 2’, 7’-dichlorfluorescein diacetate staining followed by photofluo-rography.Mitochondrial membrane potential (MMP) was examined by rhodamine 123 staining followed by photofluorogra-phy.The number of apoptotic cells was observed by Hoechst 33258 nuclear staining followed by photofluorography .RE-SULTS:After the H9c2 cells were treated with HG (35 mmol/L glucose) for 0~24 h, the protein expression of RIP3 in the H9c2 cells was significantly increased at 3 h, 6 h, 9 h, 12 h and 24 h, reaching the maximum level at 24 h.Pretreat-ment of the cells with 400μmol/L NaHS (a donor of H2S) or co-treatment of the cells with necrostatin-1 (Nec-1;a speci-fic inhibitor of necroptosis) considerably blocked the up-regulation of RIP3 protein induced by HG.Moreover, pretreatment with NaHS or co-treatment with Nec-1 obviously inhibited HG-induced injuries , leading to an increase in the cell viability , and decreases in the generation of ROS and MMP loss .On the other hand , pretreatment with NaHS also reduced the num-ber of apoptotic cells and the protein level of cleaved caspase-3 in the HG-treated H9c2 cardiomyocytes .CONCLUSION:H2 S protects H9c2 cardiomyocytes against HG-induced injury by inhibiting necroptosis .

Aim To investigate the role of ATP-sensi-tive potassium channels-Akt pathway in exogenous hy-drogen sulfide( H2 S) inhibiting the high glucose( HG)-induced injury in H9c2 cardiac cells. Methods The expression level of Akt protein was tested by Western blot assay. The cell viability was measured by cell counter kit-8(CCK-8 assay). The number of apoptotic cells was tested by Hoechst 33258 nuclear staining fol-lowed by photofluorography. The intracellular levels of reactive oxygen species ( ROS ) were detected by DCFH-DA staining followed by photofluorography. Mi-tochondrial membrane potential ( MMP ) was examined by JC-1 staining followed by photofluorography. Results H9c2 cells were treated with 35 mmol·L-1 glucose (high glucose, HG) for 0 ~24 h respectively. After treating for 3 h, the expression level of phosphorated ( p )-Akt protein began to be obviously reduced, the maximum reduced expression level was observed after the cells were exposed to HG for 24 h. Pretreatment of the cells with 50 μmol · L-1 pinacidil ( Pin, a KATP channel opener) or 400 μmol·L-1 NaHS( a donor of H2 S) prior to exposure to HG considerably blocked the down regulation of p-Akt expression level induced by HG. However, pretreatment with 1 mmol · L-1 KATP channel blocker glibenclamide( Gli) obviously attenua-ted the inhibitory effect of NaHS on HG-induced down-regulation of p-Akt expression level. On the other hand, the protective effects of NaHS against the HG-induced cardiomyocyte injury were markedly blocked by 30 μmol·L-1 Ly294002(an inhibitor of Akt), as indicated by the decrease in cell viability and MMP dissipation as well as the increases in the number of apoptotic cells and ROS generation. Conclution KATP channels-Akt pathway mediates the protective effect of H2 S against the HG-induced cardiac injury.

Aim To investigate the role of the interac-tion between necroptosis ( Nec ) and p38 mitogen-acti-vated protein kinase ( MAPK) pathway in the high glu-cose (HG)-induced H9c2 cardiac cells injury.Meth-ods The cell viability was measured by cell counter kit-8 assay .The intracellular level of reactive oxygen species ( ROS ) was tested by DCFH-DA stating fol-lowed by photofluorography .Mitochondrial membrane potential ( MMP) was detected by Rhodamine 123 stai-ning followed by photofluorography . The expression levels of receptor interaction protein 3 ( RIP3, an indi-cator of Nec ) and p38 MAPK protein were tested by Western blot assay .Results The treatment of H9c2 cardiac cells with 35 mmol? L-1 glucose ( high glu-cose, HG) for 24 h induced considerable injuries , in-cluding a decrease in cell viability , increases in ROS generation as well as MMP loss .The co-treatment of the cells with 100 μmol? L-1 necrostatin-1(Nec-1,a specific inhibitor of Nec ) and HG for 24 h or the pre-treatment of the cells with 3 μmol? L-1 SB 2 0 3 5 8 0 ( an inhibitor of p38MAPK) for 60 min before HG exposure attenuated the above injuries induced by HG .Moreo-ver, the treatment of the cells with HG for 1,3,6,9, 12 ,24 ,36 and 48 h significantly increased the expres-sion levels of RIP3, peaking at 24 h.The co-treatment of the cells with 100 μmol? L-1 Nec-1 or the pre-treatment of the cells with 3 μmol? L-1 SB203580 considerably blocked the up-regulation of RIP3 expres-sion induced by HG .On the other hand , the co-treat-ment of the cells with 100 μmol? L-1 Nec-1 alleviated the HG-induced up-regulation of the expression of p-p38MAPK.Conclusion The interaction between Nec and p38 MAPK pathway mediates the HG-induced inju-ry in H9c2 cardiac cells.

AIM:Tostudywhe ther theangiotens in-(1-7)[Ang-(1-7)]/Mas receptor axis protects cardio-myocytes against high glucose (HG)-induced injury by inhibiting nuclear factor-κB (NF-κB) pathway.METHODS:The cell viability was measured by CCK-8 assay.The intracellular levels of reactive oxygen species ( ROS) were detected by DCFH-DA staining .The number of apoptotic cells was tested by Hoechst 33258 nuclear staining .Mitochondrial membrane potential ( MMP) was examined by JC-1 staining.The levels of NF-κB p65 subunit and cleaved caspase-3 protein were de-termined by Western blotting.RESULTS: Treatment of H9c2 cardiac cells with 35 mmol/L glucose (HG) for 30, 60, 90, 120 and 150 min significantly enhanced the levels of phosphorated ( p) NF-κB p65, peaking at 60 min.Co-treatment of the cells with 1 μmol/L Ang-(1-7) and HG for 60 min attenuated the up-regulation of p-NF-κB p65 induced by HG. Co-treatment of the cells with Ang-(1-7) at concentrations of 0.1~30μmol/L and HG for 24 h inhibited HG-induced cy-totoxicity, evidenced by an increase in cell viability .On the other hand, 1 μmol/L Ang-(1-7) ameliorated HG-induced apoptosis, oxidative stress and mitochondrial damage , indicated by decreases in the number of apoptotic cells , cleaved caspase-3 level, ROS generation and MMP loss .However, the above cardioprotective effects of Ang-(1-7) were markedly blocked by A-779, an antagonist of Ang-(1-7) receptor (Mas receptor).Similarly, co-treatment of H9c2 cardiac cells with 100 μmol/L PDTC ( an inhibitor of NF-κB) and HG for 24 h also obviously reduced the above injuries induced by HG.CONCLUSION:Ang-(1-7)/Mas receptor axis prevents the cardiomyocytes from the HG-induced injury by inhibiting NF-κB pathway .