tRNA-derived fragment 3'tRF-AlaAGC modulates cell chemoresistance and M2 macrophage polarization via binding to TRADD in breast cancer.

BACKGROUND: Drug resistance, including Adriamycin-based therapeutic resistance, remains a challenge in breast cancer (BC) treatment. Studies have revealed that macrophages could play a pivotal role in mediating the chemoresistance of cancer cells. Accumulating evidence suggests that tRNA-Derived small RNAs (tDRs) are associated the physiological and pathological processes in multiple cancers. However, the underlying mechanisms of tDRs on chemoresistance of BC in tumor-associated macrophages remain largely unknown. METHODS: The high-throughput sequencing technique was used to screen tDRs expression profile in BC cells. Gain- and loss-of-function experiments and xenograft models were performed to verify the biological function of 3'tRF-Ala-AGC in BC cells. The CIBERSORT algorithm was used to investigate immune cell infiltration in BC tissues. To explore the role of 3'tRF-Ala-AGC in macrophages, M2 macrophages transfected with 3'tRF-Ala-AGC mimic or inhibitor were co-cultured with BC cells. Effects on Nuclear factor-κb (NF-κb) pathway were investigated by NF-κb nuclear translocation assay and western blot analysis. RNA pull-down assay was performed to identify 3'tRF-Ala-AGC interacting proteins. RESULTS: A 3'tRF fragment of 3'tRF-AlaAGC was screened, which is significantly overexpressed in BC specimens and Adriamycin-resistant cells. 3'tRF-AlaAGC could promote cell malignant activity and facilitate M2 polarization of macrophages in vitro and in vivo. Higher expression of M2 macrophages were more likely to have lymph node metastasis and deeper invasion in BC patients. Mechanistically, 3'tRF-AlaAGC binds Type 1-associated death domain protein (TRADD) in BC cells, and suppression of TRADD partially abolished the enhanced effect of 3'tRF-AlaAGC mimic on phenotype of M2. The NF-κb signaling pathway was activated in BC cells co-cultured with M2 macrophages transfected with 3'tRF-AlaAGC mimic. CONCLUSIONS: 3'tRF-AlaAGC might modulate macrophage polarization via binding to TRADD and increase the effect of M2 on promoting the chemoresistance in BC cells through NF-κb signaling pathway.

MiR-10b-5p Impairs TET2-Mediated Inhibition of PD-L1 Transcription Thus Promoting Immune Evasion and Tumor Progression in Glioblastoma.

Glioblastoma (GBM) is a highly aggressive primary brain tumor that shows intratumoral heterogeneity at the cellular and molecular level. Activation of programmed death receptor 1 (PD-1) interaction with its ligand PD-L1 is a well-known mechanism requisite for immune evasion deployed by malignant tumors including GBM. Herein, we set out to dissect the mechanism explaining the regulation of PD-L1 gene expression in GBM. The clinical samples consisted of 37 GBM tissues and 18 normal brain tissues. GBM cell model was treated by microRNA (miRNA) inhibitor, DNA constructs, and siRNAs. Assays of CCK-8 and Transwell insert were employed to assess the survival, migratory and invasive ability of GBM cell model. The immunosuppressive factor production, T cell apoptosis, and T cell cytotoxicity to GBM cells were evaluated in the co-culture system. GBM exhibited more miR-10b-5p abundance than normal at both tissue and cellular level. Suppression of miR-10b-5p weakened the ability of GBM cell model to survive, migrate, and invade, decreased the release of immunosuppressive factors, reduced T cell apoptosis, and strengthened the T cell cytotoxicity to GBM cell model. MiR-10b-5p conferred a negative control of Ten-eleven translocation 2 (TET2) that was downregulated in GBM. The functions of miR-10b-5p on GBM cell aggressiveness and immune evasion were mediated by TET2. TET2 recruited histone deacetylases HDAC1 and HDAC2 into the PD-L1 promoter region thus inhibiting its transcription. The study demonstrated the importance of miR-10b-5p-mediated repression of TET2 in PD-L1-driven immune evasion and their potential for immunotherapeutic targeting in GBM.

[Analysis of IVD gene variants in four children with isovalerate acidemia].

OBJECTIVE: To detect variants of IVD gene among 4 neonates with suspected isovalerate acidemia in order to provide a guidance for clinical treatment. METHODS: 111 986 newborns and 7461 hospitalized children with suspected metabolic disorders were screened for acyl carnitine by tandem mass spectrometry. Those showing a significant increase in serum isovaleryl carnitine (C5) were analyzed for urinary organic acid and variants of the IVD gene. RESULTS: Four cases of isovalerate acidemia were detected, which included 2 asymptomatic newborns (0.018‰, 2/111 986) and 2 children suspected for metabolic genetic diseases (0.268‰, 2/7461). The formers had no obvious clinical symptoms. Analysis of acyl carnitine has suggested a significant increase in C5, and urinary organic acid analysis has shown an increase in isovaleryl glycine and 3-hydroxyisovalerate. Laboratory tests of the two hospitalized children revealed high blood ammonia, hyperglycemia, decreased red blood cells, white blood cells, platelets and metabolic acidosis. The main clinical manifestations have included sweaty foot-like odor, feeding difficulty, confusion, drowsiness, and coma. Eight variants (5 types) were detected, which included c.158G>A (p.Arg53His), c.214G>A (p.Asp72Asn), c.548C>T (p.Ala183Val), c.757A>G (p.Thr253Ala) and 1208A>G (p.Tyr403Cys). Among these, c.548C>T and c.757A>G were unreported previously. None of the variants was detected by next generation sequencing of 2095 healthy newborns, and all variants were predicted to be likely pathogenic based on the guidelines from the American College of Medical Genetics and Genomics. CONCLUSION: The incidence of isovalerate acidemia in Liuzhou area is quite high. Screening of metabolic genetic diseases is therefore recommended for newborns with abnormal metabolism. The discovery of novel variants has enriched the mutational spectrum of the IVD gene.

[Analysis of CGDH gene variants and clinical features in three patients with glutaric aciduria type â ].

OBJECTIVE: To screen for potential variants of GCDH gene in 3 patients clinically diagnosed as glutaric aciduria type Ⅰ. METHODS: GCDH gene variants was detected by Sanger sequencing among the three children and their family members. RESULTS: Sanger sequencing showed that patient 1 carried compound heterozygosity variants of c.532G>A (p.Gly178Arg) and c.655G>A (p.Ala219Thr) of the GCDH gene, while his father and mother respectively carried heterozygous c.532G>A(p.Gly178Arg) and c.655G>A (p.Ala219Thr) variants. Patient 2 carried c.532G>A (p.Gly178Arg) and a novel c.1060G>T (p.Gly354Cys) compound heterozygous variant, while his father and mother respectively carried heterozygous c.532G>A (p.Gly178Arg) and c.1060G>T (p.Gly354Cys) variant. Patient 3 carried homozygous c.532G>A (p.Gly178Arg) variant of the GCDH gene, for which both of his parents were heterozygous carriers. CONCLUSION: The GCDH gene variant probably underlie the glutaric aciduria type Ⅰ among the 3 patients. Identifcation of the novel variant has enriched the spectrum of GCDH gene variants.

[Tandem mass spectrometry analysis and genetic diagnosis of neonates with fatty acid oxidation disorders in central and northern regions of Guangxi].

OBJECTIVE: To determine the incidence and mutational types of fatty acid oxidation disorders (FAOD) in central-northern region of Guangxi. METHODS: A total of 62 953 neonates were screened for FAOD during December 2012 and December 2017. Acyl-carnitine profiling of neonatal blood sample was performed by tandem mass spectrometry using dry blood spots on a filter paper. The diagnosis of FAOD was confirmed by organic acid profiling of urea and genetic testing. RESULTS: Eighteen cases of FAOD were diagnosed among the 62 953 neonates. Among these, primary carnitine deficiency (PCD) was the most common type (n=13), which was followed by short-chain acyl-CoA dehydrogenase deficiency (SCADD) (n=2), medium-chain acyl-CoA dehydrogenase deficiency (MCADD) (n=1), multiple acyl-CoA dehydrogenase deficiency (MADD) (n=1), and carnitine palmitoyltransferase II deficiency (CPT II D) (n=1). Genetic testing has revealed two previously unreported variants, i.e., c.337G to A (p.Gly113Arg) of ACADS gene and c.737G TO T (p.Gly246Val) of ETFA gene. CONCLUSION: PCD is the most common FAOD in central-northern Guangxi. Tandem mass spectrometry combined with genetic testing may facilitate early diagnosis of FAOD.

[SLC22A5 gene mutation analysis and prenatal diagnosis for a family with primary carnitine deficiency].

OBJECTIVE: To carry out mutation analysis and prenatal diagnosis for a family affected with primary carnitine deficiency. METHODS: Genomic DNA of the proband was extracted from peripheral blood sample 10 days after birth. The 10 exons and intron/exon boundaries of the SLC22A5 gene were subjected to PCR amplification and Sanger sequencing. The proband's mother was pregnant again two years after his birth. Fetal DNA was extracted from amniocytes and subjected to PCR and Sanger sequencing. RESULTS: Tandem mass spectrometric analysis of the proband revealed low level of plasma-free carnitine whilst organic acids in urine was normal. Compound heterozygous SLC22A5 mutations c.1195C>T (inherited from his father) and c.517delC (inherited from his mother) were detected in the proband. Prenatal diagnosis has detected no mutation in the fetus. The plasma-free carnitine was normal after birth. CONCLUSION: Appropriate genetic testing and prenatal diagnosis can prevent further child with carnitine deficiency. The identification of c.517delC, a novel mutation, enriched the spectrum of SLC22A5 mutations.

[Screening indices and their cut-off values for full-term neonates carrying ß-thalassemia gene].

OBJECTIVE: To investigate the screening indices and their cut-off values for full-term neonates carrying ß-thalassemia gene. METHODS: A retrospective analysis was performed for the clinical data of 1 193 full-term neonates who underwent ß-thalassemia screening (hemoglobin analysis with dried blood spots on neonatal heel blood filter paper and mutation detection of 17 ß-globin genes). A multivariate logistic regression analysis was used to investigate the association between screening indices and ß-thalassemia gene, and the receiver operating characteristic (ROC) curve was used to analyze the value of screening indices in determining the presence or absence of ß-thalassemia gene. RESULTS: Of the 1 193 neonates, 638 carried ß-thalassemia gene. Of the 1 193 neonates, 637 (53.39%) had no HbA2, among whom 310 carried ß-thalassemia gene and 327 did not carry this gene; 556 (46.61%) had HbA2, among whom 328 carried ß-thalassemia gene and 228 did not carry this gene. As for the neonates without HbA2, the ß-thalassemia gene group had a significantly lower HbA level and a significantly higher HbF level than the ß-thalassemia gene-negative group (P<0.01). As for the neonates with HbA2, the ß-thalassemia gene group had a significantly lower HbA level and significantly higher HbF and HbA2/HbA ratio than the ß-thalassemia gene-negative group (P<0.01). In the neonates without HbA2, HbA, gestational age, and HbA combined with gestational age had an area under the ROC curve (AUC) of 0.865, 0.515, and 0.870, respectively, in determining the presence or absence of ß-thalassemia gene (P<0.01), and HbA and HbA combined with gestational age had a similar AUC and a certain diagnostic value. In the neonates with HbA2, HbA, HbA2/HbA ratio, and HbA combined with HbA2/HbA ratio had an AUC of 0.943, 0.885, and 0.978, respectively, in determining the presence or absence of ß-thalassemia gene. The HbA combined with HbA2/HbA ratio had the largest AUC. In the neonates without HbA2, HbA had the largest AUC in determining the presence or absence of ß-thalassemia gene at the cut-off value of 11.6%, with a sensitivity of 85.81% and a specificity of 79.82%. In the neonates with HbA2, an HbA of 16.1%-22.0% and an HbA2/HbA ratio of >1.4 had the largest AUC in determining the presence or absence of ß-thalassemia gene, with a sensitivity of 91.38% and a specificity of 91.89%. CONCLUSIONS: HbA and HbA2/HbA ratio are effective indices for screening out full-term neonates carrying ß-thalassemia gene.

[Genetic diagnosis of 10 neonates with primary carnitine deficiency].

OBJECTIVE: To study the gene mutation profile of primary carnitine deficiency (PCD) in neonates, and to provide a theoretical basis for early diagnosis and treatment, genetic counseling, and prenatal diagnosis of PCD. METHODS: Acylcarnitine profile analysis was performed by tandem mass spectrometry using 34 167 dry blood spots on filter paper. The SLC22A5 gene was sequenced and analyzed in neonates with free carnitine (C0) levels lower than 10 µmol/L as well as their parents. RESULTS: In the acylcarnitine profile analysis, a C0 level lower than 10 µmol/L was found in 10 neonates, but C0 level was not reduced in their mothers. The 10 neonates had 10 types of mutations at 20 different sites in the SLC22A5 gene, which included 4 previously unreported mutations: c.976C>T, c.919delG, c.517delC, and c.338G>A. Bioinformatics analysis showed that the four new mutations were associated with a risk of high pathogenicity. CONCLUSIONS: Tandem mass spectrometry combined with SLC22A5 gene sequencing may be useful for the early diagnosis of PCD. Identification of new mutations enriches the SLC22A5 gene mutation profile.

Monitoring the Oxygen Dynamics of Brain Tissue In Vivo by Fast Acousto-Optic Scanning Microscopy: A Proposed Instrument.

The function of the brain neural circuit is highly dependent on oxygen supply. Imaging the precise oxygen distribution and dynamics are critical for understanding the relationship between neuronal activity and oxygen dynamics of the nearby capillaries. Here, we develop fast acousto-optic scanning two-photon microscopy. Combined with oxygen probes, such as PtP-C343, we can monitor oxygen dynamics at the submicron level by this real-time microscopy. In this fast acousto-optic scanning microscopy, an acousto-optic deflector (AOD), an inertia-less scanner, is used to scan the femtosecond laser. A cylindrical lens is used to compensate the 'cylindrical lens effect' of AOD and a prism is used to compensate the chromatic dispersion of AOD. An electro-optical modulator (EOM) and a sCMOS camera are gated to measure the phosphorescence lifetime. With a 40× water objective lens, this set-up can image a 100 µm × 100 µm field of view at a speed of 20 frames per second and a 25 µm × 8 µm field of view at a speed of 500 frames per second. This real-time two-photon microscopy is expected to be a good tool for observing and recording the precise rapid oxygen dynamics in the cerebral cortex, which will facilitate studies of oxygen metabolism in neurosciences.

[An analysis of clinical characteristics and gene mutation in two patients with medium- and short-chain acyl-CoA dehydrogenase deficiency].

Medium- and short-chain acyl-CoA dehydrogenase deficiency is a disorder of fatty acid ß-oxidation. Gene mutation prevents medium- and short-chain fatty acids from entry into mitochondria for oxidation, which leads to multiple organ dysfunction. In this study, serum acylcarnitines and the organic acid profile in urea were analyzed in two children whose clinical symptoms were hypoglycemia and metabolic acidosis. Moreover, gene mutations in the two children and their parents were evaluated. One of the patients was a 3-day-old male who was admitted to the hospital due to neonatal asphyxia, sucking weakness, and sleepiness. The serum acylcarnitine profile showed increases in medium-chain acylcarnitines (C6-C10), particularly in C8, which showed a concentration of 3.52 µmol/L (reference value: 0.02-0.2 µmol/L). The analysis of organic acids in urea gave a normal result. Sanger sequencing revealed a reported c.580A>G (p.Asn194Asp) homozygous mutation at exon 7 of the ACADM gene. The other patient was a 3-month-old female who was admitted to the hospital due to cough and recurrent fever for around 10 days. The serum acylcarnitine profile showed an increase in serum C4 level, which was 1.66 µmol/L (reference value: 0.06-0.6 µmol/L). The analysis of organic acids in urea showed an increase in the level of ethyl malonic acid, which was 55.9 (reference value: 0-6.2). Sanger sequencing revealed a reported c.625G>A (p.Gly209Ser) homozygous mutation in the ACADS gene. This study indicates that screening tests for genetic metabolic diseases are recommended for children who have unexplained metabolic acidosis and hypoglycemia. Genetic analyses of the ACADM and ACADS genes are helpful for the diagnosis of medium- and short-chain acyl-CoA dehydrogenase deficiency.

[Pancytopenia and metabolic decompensation in a neonate].

A 9-day-old male patient was admitted to the hospital because of cough, anhelation, feeding difficulty and lethargy. The diagnostic examinations indicated pulmonary infection, severe metabolic acidosis, hyperglycemia, hyperammonemia and pancytopenia in the patient. Blood and urine screening and isovaleryl-CoA dehydrogenase (IVD) gene detection for inherited metabolic diseases were performed to clarify the etiology. Tandem mass spectrometric screening for blood showed an elevated isovalerylcarnitine (C5) level. The organic acid analysis of urine by gas chromatography-mass spectrometry showed significantly increased levels in isovaleryl glycine and 3-hydroxyisovaleric acid. Homozygous mutations (c.1208A>G, p.Tyr403Cys) in the IVD gene were identified in the patient. His parents were heterozygous carriers. After the treatment with low-leucine diets and L-carnitine for 3 days, the patient showed a significant improvement in symptoms, but he died one week later. It is concluded that the neonates with pneumonia and metabolic decompensation of unknown etiology should be screened for genetic metabolic disease.

[CPT2 gene mutation analysis and prenatal diagnosis in a family with carnitine palmitoyltransferase II deficiency].

This study aimed to identify the type of carnitine palmitoyltransferase 2 (CPT2) gene mutation in the child with carnitine palmitoyltransferase II (CPT II) deficiency and her parents and to provide the genetic counseling and prenatal diagnosis for the family members. As the proband, a 3-month-old female baby was admitted to the hospital due to fever which had lasted for 8 hours. Tandem mass spectrometric analysis for blood showed an elevated plasma level of acylcarnitine, which suggested CPT II deficiency. The genomic DNA was extracted from peripheral blood of the patient and her parents. Five exon coding regions and some intron regions at the exon/intron boundaries of the CPT2 gene were analyzed by PCR and Sanger sequencing. Amniotic fluid was taken from the mother during the second trimester, and DNA was extracted to analyze the type of CPT2 gene mutation. Sanger sequencing results showed that two mutations were identified in the CPT2 gene of the proband: c.886C>T (p.R296X) and c.1148T>A (p.F383Y), which were inherited from the parents; the second child of the mother inherited the mutation of c.886C>T (p.R296X) and showed normal acylcarnitine spectrum and normal development after birth. It is concluded that the analysis of CPT2 gene mutations in the family suggested that the proband died of CPT II deficiency and that the identification of the mutations was helpful in prenatal diagnosis in the second pregnancy.

Variation of peripheral blood-based biomarkers for response of anti-PD-1 immunotherapy in non-small-cell lung cancer.

PURPOSE: Immune checkpoint inhibitors (ICIs) for non-small-cell lung cancer (NSCLC) are on the rise, but unfortunately, only a small percentage of patients benefit from them in the long term. Thus, it is crucial to identify biomarkers that can forecast the efficacy of immunotherapy. METHODS: We retrospectively studied 224 patients with NSCLC who underwent anti-PD-1 therapy. The role of biomarkers and clinical characteristics were assessed in a prognostic model. RESULTS: Only 14.3% of patients had both programmed death ligand 1 (PD-L1) and tumor mutational burden (TMB) outcomes, highlighting the need to investigate more available biomarkers. Our analysis found a correlation between histological PD-L1 TPS and hematological PD-1 expression. Analysis of hematological biomarkers revealed that elevated expression of CD4/CD8 and LYM% are positively associated with effective immunotherapy, while PD-1+ on T cells, NLR, and MLR have a negative impact. Moreover, high level of ΔCEA%, CYFRA21-1 and LDH may suggest ineffective ICIs. We also observed that disparate immunotherapy drugs didn't significantly impact prognosis. Lastly, by comparing squamous carcinoma and adenocarcinoma cohorts, ΔCEA%, CD3+PD-1+, CD4+PD-1+, and CD4/CD8 are more important in predicting the prognosis of adenocarcinoma patients, while age is more significant for squamous carcinoma patients. CONCLUSION: Our research has yielded encouraging results in identifying a correlation between immunotherapy's response and clinical characteristics, peripheral immune cell subsets, and biochemical and immunological biomarkers. The screened hematological detection panel could be used to forecast an NSCLC patient's response to anti-PD-1 immunotherapy with an accuracy rate of 76.3%, which could help customize suitable therapeutic decision-making.

Therapeutic mechanisms of modified Jiawei Juanbi decoction in early knee osteoarthritis: A multimodal analysis.

Modified Jiawei Juanbi decoction (MJD) is used for the treatment of early-stage knee osteoarthritis (KOA). Here, modified Jiawei Juanbi decoction (MJD) was employed for the treatment of early-stage knee osteoarthritis (KOA) and its mechanisms were assessed via metabonomics and network pharmacology. A total of 24 male Sprague-Dawley rats were randomly allocated into a normal control group, a model group, and an MJD group (n = 8 rats per group). Each rat group was further equally divided into two subgroups for investigation for either 14 or 28 days. A rat model of early-stage KOA was constructed and rats were treated with MJD. Effects were evaluated based on changes in knee circumference, mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL). We also analyzed histopathological changes in articular cartilage. High-resolution mass spectrometry was used to analyze the chemical profile of MJD, identifying 228 components. Using an LC-Q-TOF-MS metabonomics approach, 33 differential metabolites were identified. The relevant pathways significantly associated with MJD include arginine and proline metabolism, vitamin B6 metabolism, as well as the biosynthesis of phenylalanine, tyrosine and tryptophan. The system pharmacology paradigm revealed that MJD contains 1027 components and associates with 1637 genes, of which 862 disease genes are related to osteoarthritis. The construction of the MJD composition-target-KOA network revealed a total of 140 intersection genes. A total of 39 hub genes were identified via integration of betweenness centrality values greater than 100 using CytoHubba. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed several significantly affected signaling pathways including the HIF-1, AGE-RAGE (in diabetic complications), IL-17, rheumatoid arthritis and TNF pathways. Integrated-omics and network pharmacology approaches revealed a necessity for further detailed investigation focusing on two major targets, namely NOS2 and NOS3, along with their essential metabolite (arginine) and associated pathways (HIF-1 signaling and arginine and proline metabolism). Real-time PCR validated significantly greater downregulation of NOS2 and HIF-1ɑ in the MJD as compared to the model group. Molecular docking analysis further confirmed the binding of active MJD with key active components. Our findings elucidate the impact of MJD on relevant pathophysiological and metabolic networks relevant to KOA and assess the drug efficacy of MJD and its underlying mechanisms of action.

MVP enhances FGF21-induced ferroptosis in hepatocellular carcinoma by increasing lipid peroxidation through regulation of NOX4.

Ferroptosis is a novel, iron-dependent regulatory cell death mainly caused by an imbalance between the production and degradation of intracellular reactive oxygen species (ROS). Recently, ferroptosis induction has been considered a potential therapeutic approach for hepatocellular carcinoma (HCC). Fibroblast growth factor 21 (FGF21) is a new modulator of ferroptosis; however, the regulatory role of FGF21 in HCC ferroptosis has not been investigated. In this study, we explored the role of FGF21 and its underlying molecular mechanism in the ferroptotic death of HCC cells. We identified Major vault protein (MVP) as a target of FGF21 and revealed that knockdown of MVP inhibited the lipid peroxidation levels of HCC cells by decreasing NADPH oxidase 4 (NOX4, a major source of ROS) transcription, thereby attenuating the effect of FGF21-mediated ferroptosis. On the other hand, MVP overexpression showed the opposite results. Mechanistically, MVP binds to IRF1 and thus interferes with the interaction between IRF1 and the YAP1 promoter, leading to an increase in NOX4 transcription. Importantly, forced expression of IRF1 or downregulation of YAP1 partially reversed the effect of MVP overexpression on HCC ferroptosis. Furthermore, the results in xenograft tumor models suggested that overexpression of MVP can efficiently increase the level of lipid peroxidation in vivo. Taken together, these results provide new insights into the regulatory mechanism of ferroptosis in HCC.

Population pharmacokinetics, dosing optimization and clinical outcomes of biapenem in patients with sepsis.

Introduction: Biapenem is a carbapenem antibiotic widely used in Asia, can be used for the treatment of adults and children with infections due to susceptible bacteria. Although biapenem is utilized in the treatment of a diverse range of bacterial infections, current pharmacokinetic data in the context of septic populations remain limited. Consequently, our research aims to evaluate the pharmacokinetics and efficacy of biapenem within a septic population to optimize biapenem therapy. Methods: In this study, we characterized the pharmacokinetics of biapenem in septic patients using a population pharmacokinetic (PPK) approach. The clinical PK data to develop the PPK model were obtained from 317 septic patients admitted to Nanjing Drum Tower Hospital between 2018 and 2022. All patients were randomized to the modeling and validation cohorts at a 3:1 ratio, with PPK modeling and validation performed utilizing the NONMEM software. Results: The model found to best describe the available data was a two-compartment PPK model with first-order elimination characterized by the parameters clearance (CL), central volume (V1), peripheral volume (V2), and intercompartmental clearance (Q). A covariate analysis identified that creatinine clearance (CLCR) was a significant covariate influencing biapenem CL, while blood urea nitrogen (BUN) was a significant covariate influencing biapenem Q. Accoding to the clinical outcome analyses, 70% of the time that the free antimicrobial drug concentration exceeds the MIC (fT >MIC) is associated with favourable clinical outcomes. The PPK model was then used to perform Monte Carlo simulations to evaluate the probability of attaining 70% fT >MIC. Conclusions: A final PPK model of biapenem was established for patients with sepsis. The current daily dosage regimen of 1.2 g may insufficient to achieve 70% fT >MIC in septic patients. The dosage regimen of 600 mg every 6 h appears to be the optimal choice.

Improved Genetic Characterization of Congenital Adrenal Hyperplasia by Long-Read Sequencing Compared with Multiplex Ligation-Dependent Probe Amplification Plus Sanger Sequencing.

Genetic analysis of congenital adrenal hyperplasia (CAH) has been challenging because of high homology between CYP21A2 and its pseudogene CYP21A1P. This study aimed to evaluate the clinical utility of long-read sequencing (LRS) in diagnosis of CAH attributable to 21-hydroxylase deficiency by comparing with multiplex ligation-dependent probe amplification plus Sanger sequencing. In this retrospective study, 69 samples, including 49 probands from 47 families with high-risk of CAH, were enrolled and blindly subjected to detection of CAH by LRS. The genotype results were compared with control methods, and discordant samples were validated by additional Sanger sequencing. LRS successfully identified biallelic variants of CYP21A2 in the 39 probands diagnosed as having CAH. The remaining 10 probands were not patients with CAH. Additionally, LRS directly identified two pathogenic single-nucleotide variations (SNVs; c.293-13C/A>G and c.955C>T) in the presence of interference caused by nearby insertions/deletions (indels). The cis-trans configuration of two or more SNVs and indels identified in 18 samples was directly determined by LRS without family analysis. Eight CYP21A1P/A2 or TNXA/B deletion chimeras, composed of five subtypes, were identified; and the junction sites were precisely determined. Moreover, LRS determined the exact genotype in two probands who had three heterozygous SNVs/indels and duplication, which could not be clarified by control methods. These findings highlight that LRS could assist in more accurate genotype imputation and more precise CAH diagnosis.

Increase in age at onset of moyamoya disease in China over 25 years.

BACKGROUND: To explore whether the age at onset (AAO) of Chinese patients with moyamoya disease (MMD) increased over time due to a reduced exposure to leptospiral infection. METHODS: We performed an independent, multicenter, retrospective study based on data from patients with MMD who initially attended four tertiary hospitals in Hubei, China, from 1996 to 2020. After stratifying the year of MMD onset into five periods (1996-2000, 2001-2005, 2006-2010, 2011-2015, and 2016-2020), we analyzed the temporal trends in AAO and compared different classes of AAO (early-onset, < 20 years; intermediate-onset, 20-49 years; late-onset, ≥ 50 years) in each period. RESULTS: We included 1858 patients in this study, with 878 women and 980 men. Their median (IQR) AAO was 47 (39-55) years. The case AAO significantly increased at the rate of 0.94 years per year (r = 0.406, p < .0001), while no trend was observed in birth years through time (p = .512). The birth cohorts who grew up in the leptospirosis epidemic years was stably susceptible to MMD. The median (IQR) AAO has increased significantly from 26 (14-37) years (1996-2000) to 51 (43-57) years (2016-2020) (p < .0001). The proportion of early-onset MMD was significantly higher in 1996-2000 (33.3%, p < .0001) and 2001-2005 (10.4%, p < .001). The AAO shows an aging trend that the proportion of late-onset MMD went from 4.5% (2001-2005) to 54.5% (2016-2020) (p < .0001). CONCLUSIONS: The AAO of MMD was increasing during a recent 25-year period in China, which may reflect a birth cohort effect that resulted from environmental changes. The disparity risk of birth cohorts with MMD changed with leptospirosis epidemics, suggesting leptospiral exposure might be a potential risk factor.

Prognostic performance of the NRS2002, NUTRIC, and modified NUTRIC to identify high nutritional risk in severe acute pancreatitis patients.

Background: Acute pancreatitis (AP) is the most common gastrointestinal disease requiring hospital admission. AP patients are categorized as mild, moderately severe, and severe AP (SAP). For SAP patients, malnutrition increases susceptibility to infection and mortality. The Nutritional Risk Screening 2002 (NRS 2002), the Nutrition Risk in Critically Ill (NUTRIC) score and modified Nutrition Risk in Critically Ill (mNUTRIC) are nutritional risk screening tools of critically ill patients and have not been validated in patients with SAP. It is essential to evaluate the prognostic performance of these nutritional risk screening tools. Materials and methods: A retrospective study was designed to validate the NRS 2002, NUTRIC, and mNUTRIC when applied to SAP patients. Receiver operating characteristic curves were plotted to investigate the predictive ability of clinical outcomes by comparing areas under the curve (AUC). Appropriate cut-offs were calculated by using Youden's index. Patients were identified as being at high nutritional risk according to the calculated cut-off values. The effects of different scoring systems on mortalities were calculated using the Cox proportional hazards model. Logistic regression was used to assess the association between the energy provision and 28-day mortality. Results: From January 2013 to December 2019, 234 SAP patients were included and analyzed. Patients categorized as high nutritional risk by the NRS 2002 (12.6% versus 1.9% for 28-day and 20.5% versus 3.7% for 90-day), NUTRIC (16.2% versus 0.0% for 28-day and 27.0% versus 0.0% for 90-day), and mNUTRIC (16.4% versus 0.0% for 28-day and 26.4% versus 0.8% for 90-day) had significant higher mortality than those categorized as low nutritional risk. The NUTRIC (AUC: 0.861 for 28-day mortality and 0.871 for 90-day mortality, both cut-off value ≥3) and mNUTRIC (AUC: 0.838 for 28-day and 0.828 for 90-day mortality, both cut-off value ≥3) showed better predictive ability of the 28- and 90-day mortality than the NRS 2002 (AUC: 0.706 for 28-day mortality and 0.695 for 90-day mortality, both cut-off value ≥5). Conclusion: The NRS 2002, NUTRIC, and mNUTRIC scores were predictors for the 28- and 90-day mortalities. The NUTRIC and mNUTRIC showed better predictive ability compared with the NRS 2002 when applied to SAP patients.