Cytochrome P450 (CYP450) enzymes, as versatile biocatalysts with the broadest range of catalytic reactions in nature, play critical roles in the metabolism of medicinal plants. They are involved in various oxidative modification processes of active ingredients, facilitating both the synthesis and degradation of bioactive substances. This review delves into the classification, structure, and catalytic mechanisms of CYP450 enzymes, emphasizing their indispensable roles in plant biosynthesis. Using representative cases, including the biosynthetic pathways of tanshinones, artemisinin, celastrol, paclitaxel, and berberine, this review highlights the functional importance of specific CYP450s. For instance, CYP71AV1 catalyzes the production of artemisinin and artemisinic aldehyde, with its activity directly affecting artemisinin yield. Similarly, CYP76AH1 and CYP76AK1 play pivotal roles in the backbone construction and postmodification of tanshinones, acting as key players in their metabolic network. In the case of celastrol, CYP712K1, CYP712K2, and CYP712K3 initiate the first oxidative reaction, providing a solid foundation for subsequent biosynthetic processes. These examples highlight the pivotal role of CYP450 enzymes in the biosynthesis of medicinal plants, showcasing both their complexity and significance in plant metabolic pathways. Furthermore, this review examines the oxidative metabolism of CYP450 enzymes under aerobic conditions and their reductive metabolism in specific environments, offering deeper insights into their catalytic mechanisms. A comprehensive understanding of these processes lays the groundwork for the effective application of CYP450 enzymes in biotechnology and plant metabolic engineering.

Objective:To investigate the relationship of plasma residual cholesterol (RC), atherogenic index (AI), triglyceride (TG) to high-density lipoprotein cholesterol (HDL-C) ratio (THR) and low-density lipoprotein cholesterol (LDL-C) to HDL-C ratio (LHR) with mild cognitive impairment(MCI) in middle-aged and elderly patients with type 2 diabetes mellitus (T2DM).Methods:This was a cross-sectional study. A total of 318 T2DM patients aged 50-80 years admitted to the general practice ward of Tiantan Hospital from October 2023 to May 2024 were enrolled. The relevant clinical data were collected, including patient basic information, glycated hemoglobin, lipid test results, and cognitive level was evaluated using the Mini-Mental State Examination (MMSE) form. According to MMSE scores, the patients were divided into the MCI group ( n=102) and the normal cognitive (NC) group ( n=216). Results:Compared with the NC group, patients in MCI group had a older age, longer diabetes duration, lower education level, higher HbA1c, and higher plasma TG levels ( P<0.05). The levels of RC, AI, LHR, and THR in the MCI group were significantly higher than those in the NC group ( P<0.001). Multivariate logistic regression analysis showed that RC ( OR=1.981, P=0.045), AI ( OR=1.657, P=0.043), LHR ( OR=4.360, P=0.001), and THR ( OR=2.908, P=0.002) were independently associated with mild cognitive impairment in T2DM patients. Age-stratified analysis revealed that among T2DM patients aged 60 to 69 years, all four plasma lipid parameters were associated with the risk of MCI in T2DM patients ( P<0.05). Conclusion:Plasma levels of RC, AI, LHR and THR are significantly associated with MCI in middle-aged and elderly T2DM patients, while LHR and THR have a closer association with MCI.

Periodontal bone defects, primarily caused by periodontitis, are highly prevalent in clinical settings and manifest as bone fenestration, dehiscence, or attachment loss, presenting a significant challenge to oral health. In regenerative medicine, harnessing developmental principles for tissue repair offers promising therapeutic potential. Of particular interest is the condensation of progenitor cells, an essential event in organogenesis that has inspired clinically effective cell aggregation approaches in dental regeneration. However, the precise cellular coordination mechanisms during condensation and regeneration remain elusive. Here, taking the tooth as a model organ, we employed single-cell RNA sequencing to dissect the cellular composition and heterogeneity of human dental follicle and dental papilla, revealing a distinct Platelet-derived growth factor receptor alpha (PDGFRA) mesenchymal stem/stromal cell (MSC) population with remarkable odontogenic potential. Interestingly, a reciprocal paracrine interaction between PDGFRA⁺ dental follicle stem cells (DFSCs) and CD31⁺ Endomucin⁺ endothelial cells (ECs) was mediated by Vascular endothelial growth factor A (VEGFA) and Platelet-derived growth factor subunit BB (PDGFBB). This crosstalk not only maintains the functionality of PDGFRA⁺ DFSCs but also drives specialized angiogenesis. In vivo periodontal bone regeneration experiments further reveal that communication between PDGFRA⁺ DFSC aggregates and recipient ECs is essential for effective angiogenic-osteogenic coupling and rapid tissue repair. Collectively, our results unravel the importance of MSC-EC crosstalk mediated by the VEGFA and PDGFBB-PDGFRA reciprocal signaling in orchestrating angiogenesis and osteogenesis. These findings not only establish a framework for deciphering and promoting periodontal bone regeneration in potential clinical applications but also offer insights for future therapeutic strategies in dental or broader regenerative medicine.
Receptor, Platelet-Derived Growth Factor alpha/metabolism* ; Humans ; Neovascularization, Physiologic/physiology* ; Dental Sac/cytology* ; Single-Cell Analysis ; Transcriptome ; Mesenchymal Stem Cells/metabolism* ; Bone Regeneration ; Animals ; Dental Papilla/cytology* ; Periodontium/physiology* ; Stem Cells/metabolism* ; Regeneration ; Angiogenesis

BACKGROUND:Polyamines play a crucial role in tissue calcification.Spermidine/spermine N1-acetyltransferase 1(SAT1),as a key rate-limiting enzyme regulating intracellular polyamine metabolism,has been associated with various pathological processes.However,its role in vascular calcification remains unclear.OBJECTIVE:To investigate the role of SAT1 in rat vascular smooth muscle cell calcification.METHODS:(1)Bioinformatics analysis:Differential expression of SAT1 in human carotid atherosclerotic plaques and their surrounding healthy carotid artery tissues were using GEO datasets.PanglaoDB database was used to analyze SAT1 expression abundance and localization across different cell types through single-cell sequencing.(2)Rat vascular smooth muscle cells were divided into three groups:a control group cultured in DMEM medium,a calcification group induced by DMEM medium containing 10 mmol/L β-glycerophosphate sodium and 3 mmol/L calcium chloride,and the 50,100 μmol/L diacetylaminotriazamidine groups treated with the SAT1 inhibitor,diacetylaminotriazamidine,in addition to the calcification medium.After 7-10 days of culture,alizarin red S staining was performed,and cellular calcium content and alkaline phosphatase activity were assessed.Western blot was used to detect the protein expression of Runt-related transcription factor 2,bone morphogenetic protein 2,alpha-smooth muscle actin,and SAT1.Immunofluorescence staining was conducted to examine the expression of Runt-related transcription factor 2 and SAT1.RESULTS AND CONCLUSION:(1)Bioinformatics analysis revealed significantly upregulated expression of SAT1 and Runt-related transcription factor 2(P<0.05)in carotid atherosclerotic plaques compared with healthy carotid tissues(P<0.05).Single-cell sequencing database analysis confirmed SAT1 expression in vascular smooth muscle cells.(2)Compared with the control group,the calcification group showed significantly increased Runt-related transcription factor 2,bone morphogenetic protein 2,SAT1,calcium content,and alkaline phosphatase activity,while alpha-smooth muscle actin expression was significantly decreased(all P<0.05).Compared with the calcification group,the 50 and 100 μmol/L diacetylaminotriazamidine groups showed significantly decreased Runt-related transcription factor 2,bone morphogenetic protein 2,calcium content,and alkaline phosphatase activity,while alpha-smooth muscle actin expression was significantly increased(all P<0.05).(3)Immunofluorescence experiments demonstrated that compared with the calcification group,the expression intensity of Runt-related transcription factor 2 was significantly reduced in the 50 and 100 μmol/L diacetylaminotriazamidine groups.Overall,SAT1 may promote vascular smooth muscle cell calcification by upregulating Runt-related transcription factor 2 expression.

BACKGROUND:Polyamines play a crucial role in tissue calcification.Spermidine/spermine N1-acetyltransferase 1(SAT1),as a key rate-limiting enzyme regulating intracellular polyamine metabolism,has been associated with various pathological processes.However,its role in vascular calcification remains unclear.OBJECTIVE:To investigate the role of SAT1 in rat vascular smooth muscle cell calcification.METHODS:(1)Bioinformatics analysis:Differential expression of SAT1 in human carotid atherosclerotic plaques and their surrounding healthy carotid artery tissues were using GEO datasets.PanglaoDB database was used to analyze SAT1 expression abundance and localization across different cell types through single-cell sequencing.(2)Rat vascular smooth muscle cells were divided into three groups:a control group cultured in DMEM medium,a calcification group induced by DMEM medium containing 10 mmol/L β-glycerophosphate sodium and 3 mmol/L calcium chloride,and the 50,100 μmol/L diacetylaminotriazamidine groups treated with the SAT1 inhibitor,diacetylaminotriazamidine,in addition to the calcification medium.After 7-10 days of culture,alizarin red S staining was performed,and cellular calcium content and alkaline phosphatase activity were assessed.Western blot was used to detect the protein expression of Runt-related transcription factor 2,bone morphogenetic protein 2,alpha-smooth muscle actin,and SAT1.Immunofluorescence staining was conducted to examine the expression of Runt-related transcription factor 2 and SAT1.RESULTS AND CONCLUSION:(1)Bioinformatics analysis revealed significantly upregulated expression of SAT1 and Runt-related transcription factor 2(P<0.05)in carotid atherosclerotic plaques compared with healthy carotid tissues(P<0.05).Single-cell sequencing database analysis confirmed SAT1 expression in vascular smooth muscle cells.(2)Compared with the control group,the calcification group showed significantly increased Runt-related transcription factor 2,bone morphogenetic protein 2,SAT1,calcium content,and alkaline phosphatase activity,while alpha-smooth muscle actin expression was significantly decreased(all P<0.05).Compared with the calcification group,the 50 and 100 μmol/L diacetylaminotriazamidine groups showed significantly decreased Runt-related transcription factor 2,bone morphogenetic protein 2,calcium content,and alkaline phosphatase activity,while alpha-smooth muscle actin expression was significantly increased(all P<0.05).(3)Immunofluorescence experiments demonstrated that compared with the calcification group,the expression intensity of Runt-related transcription factor 2 was significantly reduced in the 50 and 100 μmol/L diacetylaminotriazamidine groups.Overall,SAT1 may promote vascular smooth muscle cell calcification by upregulating Runt-related transcription factor 2 expression.

Objective:To investigate the relationship of plasma residual cholesterol (RC), atherogenic index (AI), triglyceride (TG) to high-density lipoprotein cholesterol (HDL-C) ratio (THR) and low-density lipoprotein cholesterol (LDL-C) to HDL-C ratio (LHR) with mild cognitive impairment(MCI) in middle-aged and elderly patients with type 2 diabetes mellitus (T2DM).Methods:This was a cross-sectional study. A total of 318 T2DM patients aged 50-80 years admitted to the general practice ward of Tiantan Hospital from October 2023 to May 2024 were enrolled. The relevant clinical data were collected, including patient basic information, glycated hemoglobin, lipid test results, and cognitive level was evaluated using the Mini-Mental State Examination (MMSE) form. According to MMSE scores, the patients were divided into the MCI group ( n=102) and the normal cognitive (NC) group ( n=216). Results:Compared with the NC group, patients in MCI group had a older age, longer diabetes duration, lower education level, higher HbA1c, and higher plasma TG levels ( P<0.05). The levels of RC, AI, LHR, and THR in the MCI group were significantly higher than those in the NC group ( P<0.001). Multivariate logistic regression analysis showed that RC ( OR=1.981, P=0.045), AI ( OR=1.657, P=0.043), LHR ( OR=4.360, P=0.001), and THR ( OR=2.908, P=0.002) were independently associated with mild cognitive impairment in T2DM patients. Age-stratified analysis revealed that among T2DM patients aged 60 to 69 years, all four plasma lipid parameters were associated with the risk of MCI in T2DM patients ( P<0.05). Conclusion:Plasma levels of RC, AI, LHR and THR are significantly associated with MCI in middle-aged and elderly T2DM patients, while LHR and THR have a closer association with MCI.

Objective:To investigate the value of quantitative parameters derived from dual-layer spectral detector CT (SDCT) in characterizing regional lymph node (LN) status of colorectal cancer.Methods:From August 2019 to May 2020, 101 patients with colorectal cancer confirmed by pathology in the First Affiliated Hospital of Sun Yat-sen University were retrospectively collected. The largest regional LNs were matched with surgical pathology one by one and divided into metastatic LNs group (42 cases) and nonmetastatic LNs group (59 cases) according to pathological results. Based on preoperative venous phase contrast enhanced SDCT images he short-axis diameter (S) and the of the largest regional LN was measured, then its border and enhancement homogeneity were evaluated. Outlining the ROI along the edge of the LN on its widest cross section, the iodine density (ID) and effective atomic number (Z eff) were measured, then the normalized ID (nID) and normalized Z eff (nZ eff) were calculated. The χ 2 test, Fisher′s exact test, independent samples t-test or Mann-Whitney U test were used to compare the differences of each parameter between pathologically metastatic and nonmetastatic LNs and a logistic regression model was constructed. The ROC curves and area under the curve (AUC) were performed to evaluate the diagnostic performance of each parameter. DeLong test was used to compare the differences of each AUC. Results:The S, border, enhancement homogeneity, ID, Z eff, nID and nZ eff of LNs all showed significant differences between metastatic and nonmetastatic LNs (all P<0.001). The regression model constructed by S and Z eff of LNs had the highest value in differentiating metastatic and nonmetastatic LNs, with an AUC of 0.935, sensitivity and specificity of 85.7% and 89.8%, respectively. Its diagnostic value was higher than that of S, border, enhancement homogeneity (AUC 0.674-0.832, all P<0.05) and SDCT quantitative parameters (AUC 0.863-0.906, all P<0.05) of LNs. Conclusion:SDCT quantitative parameters facilitate the accurate diagnosis of regional metastatic LNs in patients with colorectal cancer, among which the multi-parameter regression model has the highest diagnostic value.

OBJECTIVETo investigate the suitable transfection condition of human epidermal cells (hECs) with human epidermal growth factor (EGF) gene by adenovirus vector (Ad-hEGF) and its effects on the biological characteristics of hECs.

METHODShECs were isolated from deprecated human fresh prepuce tissue of circumcision by enzyme digestion method and then sub-cultured. hECs of the third passage were used in the following experiments. (1) Cells were divided into non-transfection group and 5, 20, 50, 100, 150, and 200 fold transfection groups according to the random number table (the same grouping method below), with 3 wells in each group. Cells in non-transfection group were not transfected with Ad-hEGF gene, while cells in the latter six groups were transfected with Ad-hEGF gene in multiplicities of infection (MOI) of 5, 20, 50, 100, 150, and 200 respectively. The morphology of the cells was observed with inverted phase contrast microscope, and expression of green fluorescent protein of the cells was observed with inverted fluorescence microscope at transfection hour (TH) 24, 48, and 72. (2) Another three batches of cells were collected, grouped, and treated as above, respectively. Then the transfection rate of Ad-hEGF gene was detected by flow cytometer (n=3), the mass concentration of EGF in culture supernatant of cells was detected by enzyme-linked immunosorbent assay (n=6), and the proliferation activity of cells was detected by cell counting kit 8 (CCK8) and microplate reader (n=6) at TH 24, 48, and 72, respectively. (3) Cells were collected and divided into non-transfection group and transfection group, with 6 wells in each group. Cells in non-transfection group were cultured with culture supernatant of cells without transfection, while cells in transfection group were cultured with culture supernatant of cells which were transfected with Ad-hEGF gene in the optimum MOI (50). CCK8 and microplate reader were used to measure the biological activity of EGF secreted by cells on culture day 1, 3, and 5. (4) Cells were collected and divided into non-transfection group and transfection group, with 12 wells in each group. Cells in non-transfection group were not transfected with Ad-hEGF gene, while cells in transfection group were transfected with Ad-hEGF gene in the optimum MOI (50). The expression levels of cytokeratin 14 (CK14) and CK19 of cells were measured by immunofluorescence staining at TH 24. (5) Cells were collected, grouped, and treated as in (4), with 6 wells in each group. At post scratch hour (PSH) 0 (immediately after scratch), 12, 24, and 48, the migration distance of cells was observed and measured with inverted phase contrast microscope. Data were processed with analysis of variance of factorial design, analysis of variance for repeated measurement, and LSD test.

RESULTS(1) At TH 24 and 48, morphology of cells in each transfection group and non-transfection group were similar. Compared with that in non-transfection group, the cell debris increased significantly in 200 fold transfection group at TH 72. At TH 24, 48, and 72, the expression of green fluorescent protein was not seen in cells of non-transfection group, whereas it increased in cells of transfection group over transfection time. (2) The transfection rate of Ad-hEGF gene of cells in each transfection group increased gradually over transfection time. At TH 72, the transfection rates of Ad-hEGF gene of cells in 50-200 fold transfection groups were all above 90%, while the transfection rates of Ad-hEGF gene of cells in non-transfection group, 5, and 20 fold transfection groups were (0.51±0.20)%, (62.44±6.23)%, and (75.00±5.43)% respectively, which were obviously lower than the rate in 50 fold transfection group [(93.12±2.55)%, with P values below 0.01]. The mass concentration of EGF in culture supernatant of cells in each transfection group increased gradually over transfection time. At TH 72, the mass concentration of EGF in culture supernatant of cells in 50 fold transfection group was obviously higher than that in each of the other groups (with P values below 0.01). The proliferation activity of cells in each group at TH 24 and 48 was similar (with P values above 0.05). At TH 72, the proliferation activity of cells in 200 fold transfection group was obviously lower than that in other groups (with P values below 0.05). (3) On culture day 1, the biological activity of EGF secreted by cells in two groups was similar (P>0.05). On culture day 3 and 5, the biological activity of EGF secreted by cells in transfection group were obviously higher than that in non-transfection group (with P values below 0.01). (4) At TH 24, the expression levels of CK14 and CK19 of cells in transfection group were higher than those in non-transfection group. (5) The width of scratch in two groups was nearly the same at PSH 0. At PSH 12-48, the migration distance of cells in transfection group was obviously longer than that in non-transfection group (with P values below 0.01).

CONCLUSIONSThe suitable range of MOI of hECs transfected with Ad-hEGF gene is 50-150, and 50 is the optimum. hECs transfected with Ad-hEGF gene with MOI 50 can effectively express the EGF gene and keep its good abilities of proliferation, differentiation, and migration, as well.

Adenoviridae ; Cell Differentiation ; Cell Proliferation ; Cells, Cultured ; EGF Family of Proteins ; genetics ; metabolism ; Epidermis ; cytology ; Genetic Vectors ; Humans ; Keratins ; metabolism ; Male ; Transfection

To investigate the value of MRI for the diagnosis of achilles tendon closed rupture. 1.5T Maestro Class MRI scanner was used for the conventional scanning of 15 patients confirmed with achilles tendon closed ruptures by operation, and then the findings by imaging were compared with those by operation. The 15 patients proved with achilles tendon closed rupture, including 4 cases of incomplete rupture and 11 cases of complete rupture. MRI could display clearly the changes in morphology and signal of incomplete or complete closed ruptures of Achilles tendon, and the results were consistent with those by operation. MRI can make an accurate display of the lo-cation and extent of achilles tendon rupture.

Objective To explore the advantages of Argus method by comparing the accuracy and timeliness of Argus and artificial methods for measuring femoral head necrosis area in MRI scanning.Methods Totally 17 patients (31 hips) were measured with Argus and artificial methods respectively for the necrosis area, and then the measuring results and time were compared, and the correlation was investigated between the results and the patients' pain degree, along with that between the results and the extent of femoral head collapse.Results The necrosis area ratios determined by Argus and artificial methods were (33.5±4.08)%and (34.6±4.06)%respectively, with no statistical difference between the ratios (P>0.05). The time consumed by artificial method was (21.3 ±3.62)min, significantly longer than (7.89 ±1.03)min by Argus method, with P<0.001. Regression analysis proved that the necrosis areas were positively correlated with the patients' pain degree, and the correlation coefficient by Argus method was 0.807 8, more than 0.740 9 by artificial method. The femoral heads of 11 cases(16 hips) collapsed in the follow-up period, the necrosis areas were positively correlated with the patients collapse level, but the correlation coefficient by Argus method was 0.783 8, more than 0.726 7 by artificial method.Conclusion Argus method gains high accuracy and timeliness when used in MRI scanning of femoral head necrosis area, and thus is worth popularizing clinically.