G protein-coupled receptors (GPCRs) are significant drug targets, but their potential in cancer therapy remains underexplored. Conventional GPCR agonists or antagonists have shown limited effectiveness in cancer treatment, necessitating new GPCR-targeting strategies for more effective therapies. This study discovers that Yersinia pestis LcrV, a crucial linker protein for plague infection, acts as a biased agonist of a GPCR, the formyl peptide receptor 1 (FPR1). The LcrV protein induces unique conformational changes in FPR1, resulting in G proteins being activated in a distinctive state without subunit dissociation. This leads to a biased signaling profile characterized by cyclic adenosine monophosphate (cAMP) responses and β-arrestin2 recruitment, but not calcium mobilization. In FPR1-expressing triple-negative breast cancer (TNBC) cells, LcrV bi-directionally modulates intracellular signaling pathways, downregulating extracellular signal-regulated kinases (ERK1/2) and Akt pathways while upregulating Jun N-terminal kinase (JNK) and p38 pathways. This dual modulation results in cell cycle arrest and the inhibition of TNBC cell proliferation. In TNBC xenograft mouse models, long-term LcrV treatment inhibits tumor growth more effectively than a conventional FPR1 antagonist. Additionally, LcrV treatment reprograms tumor cells by reducing stemness-associated proteins OCT4 and c-MYC. Our findings highlight the potential of biased GPCR agonists as a novel GPCR-targeting strategy for cancer treatment.

BACKGROUND:Skeletal muscle insulin resistance is the key pathological link of type 2 diabetes.The traditional Chinese medicine compound Roujishuncuiyin can effectively improve skeletal muscle insulin resistance,but its mechanism has not been clarified.OBJECTIVE:To explore the mechanism of Roujishuncuiyin on skeletal muscle insulin resistance in type 2 diabetes mice.METHODS:Forty db/db mice with type 2 diabetes mellitus were randomized into a model group,a low-dose Roujishuncuiyin group,a high-dose Roujishuncuiyin group,and a positive drug group,with 10 mice in each group.The latter three administration groups were given 157.5 mg/g and 630 mg/g Roujishuncuiyin and 200 mg/g metformin hydrochloride aqueous solution by gavage once a day,respectively.In addition,10 db/dm mice were selected as the blank control group.Mice in the model and blank control groups were given the same dose of 0.9％NaCl solution by gavage.After 12 weeks of intervention,fasting blood glucose was measured in each group of mice,and oral glucose tolerance test was performed to calculate the area under the blood glucose curve.ELISA was used to detect serum insulin level and calculate the resistance index.Mitochondrial structure of skeletal muscle tissue was observed under transmission electron microscopy.Western blot was used to detect the expression levels and phosphorylation levels of protein kinase B(AKT)and glycogen synthase kinase 3β(GSK-3β)proteins in skeletal muscle.RESULTS AND CONCLUSION:(1)Compared with the blank control group,fasting blood glucose,fasting insulin and insulin resistance index were significantly higher in the model group(P＜0.05),the area under the curve of the oral glucose tolerance test was significantly increased(P＜0.05),the expression of p-AKT and p-GSK3β proteins in tibialis anterior muscle was significantly decreased(P＜0.05),and there was a large amount of mitochondrial damage in tibialis anterior muscle and a large number of lipid droplets in the interstitium.(2)Compared with the model group,fasting blood glucose,fasting insulin,and insulin resistance index were significantly reduced in the low-and high-dose Roujishuncuiyin groups and the positive control group(P＜0.05),the area under the curve of the oral glucose tolerance test was reduced(P＜0.05),the expression of p-AKT and p-GSK3β proteins in the tibialis anterior muscle was significantly elevated(P＜0.05),and mitochondrial damage in the tibialis anterior muscle was significantly ameliorated,with decreased lipid droplets in the interstitium.(3)The above indexes were better in the high-dose Roujishuncuiyin group than the low-dose Roujishuncuiyin group(P＜0.05),while there was no significant difference between the high-dose Roujishuncuiyin group and positive control group(P＞0.05).To conclude,by upregulating the protein levels of p-AKT and p-GSK3β in skeletal muscle tissue,the traditional Chinese medicine compound Roujishuncuiyin can improve structural disorders and mitochondrial morphology in skeletal muscle tissue,reduce insulin resistance in the skeletal muscle and regulate glucose homeostasis in the body.

BACKGROUND:Skeletal muscle insulin resistance is the key pathological link of type 2 diabetes.The traditional Chinese medicine compound Roujishuncuiyin can effectively improve skeletal muscle insulin resistance,but its mechanism has not been clarified.OBJECTIVE:To explore the mechanism of Roujishuncuiyin on skeletal muscle insulin resistance in type 2 diabetes mice.METHODS:Forty db/db mice with type 2 diabetes mellitus were randomized into a model group,a low-dose Roujishuncuiyin group,a high-dose Roujishuncuiyin group,and a positive drug group,with 10 mice in each group.The latter three administration groups were given 157.5 mg/g and 630 mg/g Roujishuncuiyin and 200 mg/g metformin hydrochloride aqueous solution by gavage once a day,respectively.In addition,10 db/dm mice were selected as the blank control group.Mice in the model and blank control groups were given the same dose of 0.9％NaCl solution by gavage.After 12 weeks of intervention,fasting blood glucose was measured in each group of mice,and oral glucose tolerance test was performed to calculate the area under the blood glucose curve.ELISA was used to detect serum insulin level and calculate the resistance index.Mitochondrial structure of skeletal muscle tissue was observed under transmission electron microscopy.Western blot was used to detect the expression levels and phosphorylation levels of protein kinase B(AKT)and glycogen synthase kinase 3β(GSK-3β)proteins in skeletal muscle.RESULTS AND CONCLUSION:(1)Compared with the blank control group,fasting blood glucose,fasting insulin and insulin resistance index were significantly higher in the model group(P＜0.05),the area under the curve of the oral glucose tolerance test was significantly increased(P＜0.05),the expression of p-AKT and p-GSK3β proteins in tibialis anterior muscle was significantly decreased(P＜0.05),and there was a large amount of mitochondrial damage in tibialis anterior muscle and a large number of lipid droplets in the interstitium.(2)Compared with the model group,fasting blood glucose,fasting insulin,and insulin resistance index were significantly reduced in the low-and high-dose Roujishuncuiyin groups and the positive control group(P＜0.05),the area under the curve of the oral glucose tolerance test was reduced(P＜0.05),the expression of p-AKT and p-GSK3β proteins in the tibialis anterior muscle was significantly elevated(P＜0.05),and mitochondrial damage in the tibialis anterior muscle was significantly ameliorated,with decreased lipid droplets in the interstitium.(3)The above indexes were better in the high-dose Roujishuncuiyin group than the low-dose Roujishuncuiyin group(P＜0.05),while there was no significant difference between the high-dose Roujishuncuiyin group and positive control group(P＞0.05).To conclude,by upregulating the protein levels of p-AKT and p-GSK3β in skeletal muscle tissue,the traditional Chinese medicine compound Roujishuncuiyin can improve structural disorders and mitochondrial morphology in skeletal muscle tissue,reduce insulin resistance in the skeletal muscle and regulate glucose homeostasis in the body.

Objective:To analyze regional differences in MMACHC gene variations among patients with cblC-type methylmalonic acidemia (MMA) in China and to explore the relationship between these variations and neonatal screening, biochemical markers and prognosis.Methods:Retrospective case summary. Clinical and laboratory data, including general condition, biochemical markers and genetic analysis, were collected from 1 859 cblC MMA patients from 2005 to 2023. Patients were divided into 7 groups according to their regions: north China, northeast China, east China, central China, south China, southwest China and northwest China. They were also classified into neonatal screening and non-neonatal screening groups. Mann-Whitney U and Kruskal-Wallis tests were used to compare biochemical marker levels. In contrast, the Chi-square test was applied to compare MMACHC gene variant frequencies, neonatal screening proportion, onset age and prognosis between groups. Results:Among 1 859 cases of cblC MMA, 1 019 were male and 840 were female, with a consultation age of 1.0 (0.1, 5.0) month. A total of 1 787 cases carried compound heterozygous or homozygous variants and only 1 variant site was identified in 72 cases. The 10 most frequent variants were c.609G>A (1 238 cases), c.658_660delAAG (343 cases), c.80A>G (284 cases), c.482G>A (239 cases), c.567dupT (191 cases), c.656_658delAGA (131 cases), c.217C>T (109 cases), c.394C>T (105 cases), c.445_446delTG (51 cases) and c.1A>G (50 cases). The frequency of the c.609G>A was the lowest in northwest China (28.8% (44/154), χ2=-18.42, P<0.05). The frequency of the c.567dupT was the most common in southwest China (25.0% (20/80), χ2=71.70, P<0.001) and c.656_658delAGA had the highest frequency in northeast China (9.3% (19/205), χ2=32.08, P<0.001). Non-missense variants (91.2% (62/68), 88.5% (46/52)) and early-onset patients (90.0% (36/40), 94.4% (34/36)) were both more prevalent in southwest and south China ( χ2=14.95, 31.69, both P<0.05). The proportion of neonatal screening was the lowest in south China (22.2% (8/36), χ2=98.48, P<0.05), where the mortality rate was the highest (19.1% (4/21), χ2=38.98, P<0.001). East China exhibited the highest frequency of missense variants (21.5% (339/1 579)), the highest proportion of patients identified through neonatal screening (54.5% (465/853)), and a more significant proportion of patients with good prognosis (36.6% (227/621), χ2=14.57, 93.49, 38.98, all P<0.05). In addition, the c.482G>A variant was more frequent in patients diagnosed by neonatal screening compared to those diagnosed by other methods (8.3% (132/1 586) vs. 5.9% (122/2 060), χ2=7.97, P<0.05). Conclusions:The frequency of MMACHC gene variation varies across different regions. The c.609G>A was least frequent in northwest China, c.567dupT was most common in southwest China, and c.656_658delAGA was most prevalent in northeast China. South China had the lowest neonatal screening rate and the highest mortality. At the same time, east China exhibited the highest frequency of missense variants, the highest proportion of patients identified through neonatal screening and the best prognosis. The c.482G>A variant was more frequent in patients diagnosed by neonatal screening compared to those diagnosed by other methods.

This article reports a patient with extensive high-pressure injection injury of the hand combined with deep chemical burn caused by high-pressure injection of industrial cement materials was diagnosed and treated in the Department of Hand Surgery, Xiaolan Hospital Affiliated to Southern Medical University in 2022. The nerves, tendons and blood vessels of the left hand were involved, and the ulnar skin of the left thumb was extensively necrosis, and a large number of extensive cement foreign bodies remained under the skin. Part of the cement was inserted into the joint capsule of the interphalangeal joint. After emergency surgical treatment, the patient was saved successfully, and the wound healed well without chemical poisoning and other related complications, which created conditions for the second stage of flap repair.

This article reports a patient with extensive high-pressure injection injury of the hand combined with deep chemical burn caused by high-pressure injection of industrial cement materials was diagnosed and treated in the Department of Hand Surgery, Xiaolan Hospital Affiliated to Southern Medical University in 2022. The nerves, tendons and blood vessels of the left hand were involved, and the ulnar skin of the left thumb was extensively necrosis, and a large number of extensive cement foreign bodies remained under the skin. Part of the cement was inserted into the joint capsule of the interphalangeal joint. After emergency surgical treatment, the patient was saved successfully, and the wound healed well without chemical poisoning and other related complications, which created conditions for the second stage of flap repair.

In order to obtain a recombinant fusion protein of porcine β-defensin-2 and interferon αwith broad-spectrum antibacterial and antiviral activities at the same time,the genes encoding both proteins were fused in series and inserted into the pPICZαA vector.This construct was then elec-trotransferred into Pichia pastoris KM71H cells to construct an engineered strain.Following methanol-induced expression,the recombinant protein was concentrated and isolated.The cytotox-icity of the protein was assessed using the MTT assay and a porcine red blood cell hemolysis test.Subsequently,the in vitro antibacterial and antiviral activities of the recombinant fusion protein were evaluated.The results showed that the engineered strain pPICZαA-PBD2-IFNα-KM71H of Pichia pastoris was successfully obtained,and the fusion protein PBD2-IFN-α was obtained by concentrating and purifying the fermentation broth after fermentation induction and expression.Its concentration was 1.116 g/L and molecular weight was 25 kDa.When the concentration of fusion protein was lower than 4-4 g/L,it had no obvious toxicity to PK-15 cells and porcine red blood cells.The diameter of the inhibition zone produced by the fusion protein on the mixed plate of Escherichia coli and Staphylococcus aureus was(15.0±0.9)mm,which had obvious antibacterial activity.The antiviral activity of the fusion protein against VSV in PK-15 cells was 8.89 × 105 U/mL measured by Reed-Muench method.This study laid a theoretical foundation for further develo-ping the recombinant fusion protein as an antibacterial and antiviral product.

In order to obtain a recombinant fusion protein of porcine β-defensin-2 and interferon αwith broad-spectrum antibacterial and antiviral activities at the same time,the genes encoding both proteins were fused in series and inserted into the pPICZαA vector.This construct was then elec-trotransferred into Pichia pastoris KM71H cells to construct an engineered strain.Following methanol-induced expression,the recombinant protein was concentrated and isolated.The cytotox-icity of the protein was assessed using the MTT assay and a porcine red blood cell hemolysis test.Subsequently,the in vitro antibacterial and antiviral activities of the recombinant fusion protein were evaluated.The results showed that the engineered strain pPICZαA-PBD2-IFNα-KM71H of Pichia pastoris was successfully obtained,and the fusion protein PBD2-IFN-α was obtained by concentrating and purifying the fermentation broth after fermentation induction and expression.Its concentration was 1.116 g/L and molecular weight was 25 kDa.When the concentration of fusion protein was lower than 4-4 g/L,it had no obvious toxicity to PK-15 cells and porcine red blood cells.The diameter of the inhibition zone produced by the fusion protein on the mixed plate of Escherichia coli and Staphylococcus aureus was(15.0±0.9)mm,which had obvious antibacterial activity.The antiviral activity of the fusion protein against VSV in PK-15 cells was 8.89 × 105 U/mL measured by Reed-Muench method.This study laid a theoretical foundation for further develo-ping the recombinant fusion protein as an antibacterial and antiviral product.

OBJECTIVE: To explore the disease spectrum for abnormal 3-hydroxyisovalerylcarnitine (C5OH) metabolism identified through newborn screening and clinical diagnosis patients and the key points for differential diagnosis so as to raise the awareness of pediatricians for such diseases. METHODS: Clinical data of 85 neonates with abnormal C5OH metabolism identified from February 2004 to January 2022 at Xinhua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine were collected. Their clinical manifestations and results of tandem mass spectrometry (MS/MS), gas chromatography mass spectrometry (GC-MS) and genetic testing were retrospectively analyzed. RESULTS: Among the 85 cases, 46 (54.1%) were identified by neonate screening, whilst 39 (45.9%) were clinically diagnosed patients. Five diseases were diagnosed, including 28 cases with multiple carboxylase deficiency (MCD, 32.9%), 29 cases with 3-methylcrotonyl-coenzymeAcarboxylasedeficiency (MCCD, 34.1%), 4 cases with 3-methylglutaconic acid (3-MGA, 4.7%), 7 cases with 3-hydroxy-3-methylglutaric acid (3-HMG, 8.2%), and 17 cases with beta-ketothiolase deficiency (BKD, 20.0%). The disorders were characterized by sudden onset, anorexia, vomiting, diarrhea, abnormal breathing, consciousness disorder, spasm and developmental delay. CONCLUSION Among newborns with abnormal C5OH metabolism, MCCD is the most common disorder, which was followed by BKD and MCD. For patients with abnormal C5OH metabolism, MCD is the most common, followed by BKD and 3-HMG. C5OH related diseases have great heterogeneity. Combination of blood acylcarnitine levels, urinary organic acid levels and genetic testing based on clinical characteristics can help to attain the diagnosis.
Humans ; Infant, Newborn ; China ; Neonatal Screening ; Retrospective Studies ; Tandem Mass Spectrometry/methods*

Abstract OBJECTIVE To quickly qualitatively analyze the chemical components in Sanzi powder compound prescription by HPLC-Q-Exactive-MS/MS. METHODS SHIMADZU GIST C18 column(4.6 mm×150 mm, 5 μm) was used, using 0.1% formic acid water-methanol as mobile phase with gradient elution, the flow rate was 0.5 mL·min-1, the column temperature was 30 ℃. Under positive and negative ion modes, the primary and secondary mass spectrometry information of Sanzi powder was scanned. Qualitative attribution of each component in Sanzi powder was carried out based on the mass spectrometry information, molecular formula, and retention time of molecular ion peaks and fragment ions analyzed using total ion flow diagrams, combined with the molecular formula and structural formula searched in the Chemspider database and references. RESULTS Based on the analysis of the mass spectrometry cleavage rules and references of various components, 95 possible chemical components were preliminarily inferred, including 39 phenolic acids, 20 tannins, 9 organic acid esters, 5 monoterpenoids, 12 iridoids, 8 triterpenes and 2 flavonoids. Among them, 57 components were from Chebulae Fructus, 30 were from Gardeniae Fructus, 10 were from Toosendan Fructus, and among these compounds, rutin from Chebulae Fructus, Gardeniae Fructus and Toosendan Fructus. CONCLUSION The HPLC-Q-Exactive-MS/MS detection method has good separation and high sensitivity, and can quickly and efficiently infer various components in Sanzi powder. It establishes a fast and efficient analytical method for identifying the chemical components in Sanzi powder.