Navigating the pitfalls of mapping DNA and RNA modifications.

Chemical modifications to nucleic acids occur across the kingdoms of life and carry important regulatory information. Reliable high-resolution mapping of these modifications is the foundation of functional and mechanistic studies, and recent methodological advances based on next-generation sequencing and long-read sequencing platforms are critical to achieving this aim. However, mapping technologies may have limitations that sometimes lead to inconsistent results. Some of these limitations are technical in nature and specific to certain types of technology. Here, however, we focus on common (yet not always widely recognized) pitfalls that are shared among frequently used mapping technologies and discuss strategies to help technology developers and users mitigate their effects. Although the emphasis is primarily on DNA modifications, RNA modifications are also discussed.

mEnrich-seq: methylation-guided enrichment sequencing of bacterial taxa of interest from microbiome.

Metagenomics has enabled the comprehensive study of microbiomes. However, many applications would benefit from a method that sequences specific bacterial taxa of interest, but not most background taxa. We developed mEnrich-seq (in which 'm' stands for methylation and seq for sequencing) for enriching taxa of interest from metagenomic DNA before sequencing. The core idea is to exploit the self versus nonself differentiation by natural bacterial DNA methylation and rationally choose methylation-sensitive restriction enzymes, individually or in combination, to deplete host and background taxa while enriching targeted taxa. This idea is integrated with library preparation procedures and applied in several applications to enrich (up to 117-fold) pathogenic or beneficial bacteria from human urine and fecal samples, including species that are hard to culture or of low abundance. We assessed 4,601 bacterial strains with mapped methylomes so far and showed broad applicability of mEnrich-seq. mEnrich-seq provides microbiome researchers with a versatile and cost-effective approach for selective sequencing of diverse taxa of interest.

Deciphering bacterial epigenomes using modern sequencing technologies.

Prokaryotic DNA contains three types of methylation: N6-methyladenine, N4-methylcytosine and 5-methylcytosine. The lack of tools to analyse the frequency and distribution of methylated residues in bacterial genomes has prevented a full understanding of their functions. Now, advances in DNA sequencing technology, including single-molecule, real-time sequencing and nanopore-based sequencing, have provided new opportunities for systematic detection of all three forms of methylated DNA at a genome-wide scale and offer unprecedented opportunities for achieving a more complete understanding of bacterial epigenomes. Indeed, as the number of mapped bacterial methylomes approaches 2,000, increasing evidence supports roles for methylation in regulation of gene expression, virulence and pathogen-host interactions.

Analytical challenges in omics research on asthma and allergy: A National Institute of Allergy and Infectious Diseases workshop.

Studies of asthma and allergy are generating increasing volumes of omics data for analysis and interpretation. The National Institute of Allergy and Infectious Diseases (NIAID) assembled a workshop comprising investigators studying asthma and allergic diseases using omics approaches, omics investigators from outside the field, and NIAID medical and scientific officers to discuss the following areas in asthma and allergy research: genomics, epigenomics, transcriptomics, microbiomics, metabolomics, proteomics, lipidomics, integrative omics, systems biology, and causal inference. Current states of the art, present challenges, novel and emerging strategies, and priorities for progress were presented and discussed for each area. This workshop report summarizes the major points and conclusions from this NIAID workshop. As a group, the investigators underscored the imperatives for rigorous analytic frameworks, integration of different omics data types, cross-disciplinary interaction, strategies for overcoming current limitations, and the overarching goal to improve scientific understanding and care of asthma and allergic diseases.

Discovering multiple types of DNA methylation from bacteria and microbiome using nanopore sequencing.

Bacterial DNA methylation occurs at diverse sequence contexts and plays important functional roles in cellular defense and gene regulation. Existing methods for detecting DNA modification from nanopore sequencing data do not effectively support de novo study of unknown bacterial methylomes. In this work, we observed that a nanopore sequencing signal displays complex heterogeneity across methylation events of the same type. To enable nanopore sequencing for broadly applicable methylation discovery, we generated a training dataset from an assortment of bacterial species and developed a method, named nanodisco ( https://github.com/fanglab/nanodisco ), that couples the identification and fine mapping of the three forms of methylation into a multi-label classification framework. We applied it to individual bacteria and the mouse gut microbiome for reliable methylation discovery. In addition, we demonstrated the use of DNA methylation for binning metagenomic contigs, associating mobile genetic elements with their host genomes and identifying misassembled metagenomic contigs.

High-content analysis identified synergistic drug interactions between INK128, an mTOR inhibitor, and HDAC inhibitors in a non-small cell lung cancer cell line.

BACKGROUND: The development of drug resistance is a major cause of cancer therapy failures. To inhibit drug resistance, multiple drugs are often treated together as a combinatorial therapy. In particular, synergistic drug combinations, which kill cancer cells at a lower concentration, guarantee a better prognosis and fewer side effects in cancer patients. Many studies have sought out synergistic combinations by small-scale function-based targeted growth assays or large-scale nontargeted growth assays, but their discoveries are always challenging due to technical problems such as a large number of possible test combinations. METHODS: To address this issue, we carried out a medium-scale optical drug synergy screening in a non-small cell lung cancer cell line and further investigated individual drug interactions in combination drug responses by high-content image analysis. Optical high-content analysis of cellular responses has recently attracted much interest in the field of drug discovery, functional genomics, and toxicology. Here, we adopted a similar approach to study combinatorial drug responses. RESULTS: By examining all possible combinations of 12 drug compounds in 6 different drug classes, such as mTOR inhibitors, HDAC inhibitors, HSP90 inhibitors, MT inhibitors, DNA inhibitors, and proteasome inhibitors, we successfully identified synergism between INK128, an mTOR inhibitor, and HDAC inhibitors, which has also been reported elsewhere. Our high-content analysis further showed that HDAC inhibitors, HSP90 inhibitors, and proteasome inhibitors played a dominant role in combinatorial drug responses when they were mixed with MT inhibitors, DNA inhibitors, or mTOR inhibitors, suggesting that recessive drugs could be less prioritized as components of multidrug cocktails. CONCLUSIONS: In conclusion, our optical drug screening platform efficiently identified synergistic drug combinations in a non-small cell lung cancer cell line, and our high-content analysis further revealed how individual drugs in the drug mix interact with each other to generate combinatorial drug response.

Endovascular Strategies and Outcomes for Aberrant Splenic Artery Aneurysms.

OBJECTIVES: Aberrant splenic artery aneurysms (ASAAs) located at the splenomesenteric trunk (SMT) and the celiacomesenteric trunk have a close anatomical relationship with the superior mesenteric artery (SMA). The aim of this study was to review our institutional experience of endovascular treatment for ASAAs and evaluate the long-term outcomes. METHODS: A retrospective review of patients with ASAAs who underwent endovascular treatment between December 2006 and December 2022 was performed. The demographics of the patients, aneurysm characteristics, treatment strategies, perioperative and long-term outcomes, and complications were analyzed. RESULTS: A total of 29 patients with ASAAs were endovascularly treated at our institution. The SMT variant occurred in the majority of the patients. All ASAAs were characterized by eccentric growth and extremely short inflow arteries. Only 1 patient's inflow artery of the aneurysm exceeded 1 cm in length. Thirteen patients were treated by coil embolization alone. Four patients received bare stent-assisted coil embolization. A combination of coil embolization and covered stent placement across the orifice of the aberrant splenic artery was performed in the remaining 12 cases. Coil migration into the SMA occurred in 2 patients during the operation. Technical success was achieved in all patients. With a median duration of 63 (34-101) months of follow-up, no intestinal ischemia, aneurysm-related death, aneurysm rupture, or sac enlargement occurred. Three cases of aneurysm sac reperfusion were observed, and 1 patient underwent reintervention with secondary embolization. Asymptomatic occlusion of the covered stent was detected in 1 patient at 2 years. CONCLUSIONS: Endovascular treatment is a safe, effective, and durable option for ASAAs. Inflow embolization might be difficult to achieve in ASAAs and poses a high risk of coil migration into the SMA. Long-term observation indicates that reasonable use of the covered stent could achieve reliable inflow artery exclusion in ASAAs without intestinal complications. CLINICAL IMPACT: Aberrant splenic artery aneurysm (ASAA) is an extremely rare entity. This study reported a large sample size of ASAAs treated by endovascular techniques with long-term follow-up. The ASAA was characterized by an extremely short inflow artery and a close anatomical relationship with the superior mesenteric artery (SMA). Endovascular treatment is a safe, effective, and durable option for ASAAs. Inflow embolization might be difficult to achieve in ASAAs and pose a high risk of coil migration into the SMA. Long-term observation indicates that reasonable use of the covered stent could achieve reliable inflow artery exclusion in ASAAs without intestinal complications.

Outcomes of Excimer Laser Ablation Combined with Drug-coated Balloon in Atherosclerotic Lesions of the Popliteal Artery.

BACKGROUND: The treatment of atherosclerotic lesions in the popliteal artery is challenging. This study aims to investigate the efficacy and safety of excimer laser ablation (ELA) combined with drug-coated balloon (DCB) for these lesions. METHODS: From June 2019 to December 2021, data of patients who underwent ELA combined with DCB in the popliteal artery were retrospectively reviewed. Demographics, lesion characteristics, periprocedural complications, and follow-up information were analyzed. The primary endpoint was primary patency. Secondary endpoints included major amputation-free survival rate, technical success, bailout stenting, clinically-driven target lesion reintervention, improvement of ankle-brachial index (ABI), and Rutherford class. RESULTS: A total of 61 patients were enrolled. The mean age was 73.4 ± 11.7 years. 20 (32.8%) patients had stenotic lesions, while 41 (67.2%) patients had chronic total occlusions. The mean length of these lesions was 7.3 ± 2.8 cm. Procedure technical success rate was 95.1%. Bailout stent was performed in 3 (4.9%) patients. Intraprocedural distal embolization occurred in 3 (4.9%) patients, while flow limiting dissections occurred in 3 (4.9%) patients. The mean ABI was significantly improved from 0.45 ± 0.13 at baseline to 0.90 ± 0.12 after ELA, 0.88 ± 0.11 at 6 months and 0.85 ± 0.12 at 12 months during the follow-up period. The median follow-up time was 28.2 ± 6.1 months. Reintervention was performed in 5 (8.2%) patients. The 2-year primary patency was 83.5%. CONCLUSIONS: ELA combined with DCB is a safe and effective strategy in the treatment of popliteal artery atherosclerotic lesions with low rates of bail-out stenting and high primary patency.

The impacts of national centralized drug procurement policy on drug utilization of medical institutions: an empirical study in a county-level hospital in China.

PURPOSE: Under the background of the regular implementation of the National Centralized Drug Procurement (NCDP) policy, this study aimed to assess the impacts of the NCDP policy on drug utilization of county-level medical institutions, and probe into the influencing factors of the changes in drug utilization. METHOD: A pre-post study was applied using inpatient data from a county-level medical institution in Nanjing. Drug utilization behavior of medical institutions of 88 most commonly used policy-related drugs (by generic name, including bid-winning and bid-non-winning brands) was analyzed, and the substitution of bid-winning brands for brand-name drugs after policy intervention was evaluated. RESULTS: After policy intervention, 43.18% of policy-related drugs realized the substitution of bid-winning brands for bid-non-winning brands (6.82% of complete substitution, 36.36% of partial substitution). Meanwhile, 40.90% of policy-related drugs failed to realize brand substitution. Multiple factors affected brand substitution, including: (1) Policy effect: brand substitution was more obvious after the intervention of the first and third round of NCDP. (2) Drug market competition: the greater the price reduction of bid-non-winning brands, the more the drugs for the same indication, the more likely that medical institutions keep using the same brands as they did before policy intervention. (3) Previous drug utilization of medical institutions: brand substitution was more obvious in drugs with large number of prescriptions and weak preference for brand-name drugs. CONCLUSION: The NCDP policy promoted the substitution of bid-winning brands for bid-non-winning brands. However, the NCDP policy remained to be further implemented in county-level medical institutions. Policy implememtation efforts, drug market competition and drug utilization of medical institutions would affect the implementation of the NCDP policy.

Research on a Corrosion Detection Method for Oil Tank Bottoms Based on Acoustic Emission Technology.

This paper presents an acoustic emission (AE) detection method for refined oil storage tanks which is aimed towards specialized places such as oil storage tanks with high explosion-proof requirements, such as cave oil tanks and buried oil tanks. The method utilizes an explosion-proof acoustic emission instrument to detect the floor of a refined oil storage tank. By calculating the time difference between the defective acoustic signal and the speed of acoustic wave transmission, a mathematical model is constructed to analyze the detected signals. An independent channel AE detection system is designed, which can store the collected data in a piece of independent explosion-proof equipment, and can analyze and process the data in a safe area after the detection, solving the problems of a short signal acquisition distance and the weak safety protection applied to traditional AE instruments. A location analysis of the AE sources is conducted on the bottom plate of the tank, evaluating its corrosion condition accurately. The consistency between the evaluation and subsequent open-tank tests confirms that using AE technology effectively captures corrosion signals from oil storage tanks' bottoms. The feasibility of carrying out online inspection under the condition of oil storage in vertical steel oil tanks was verified through a comparison with open inspections, which provided a guide for determining the inspection target and opening order of large-scale oil tanks.

Engineering design and application of large-scale oil-based drilling cuttings treatment project.

Thermal phase separation technology is a new comprehensive treatment technology, which heats oil-based cuttings to a certain temperature to vaporize oil and water components. Based on a large oil-based drilling cuttings comprehensive utilization project, the engineering design and application effect of thermal phase separation technology were analysed. The practice shows that thermal phase separation technology can reduce the oil content of purified residue to 0.1-0.2%, the average recovery rate of base oil is 94.12% and the annual recovery of base oil is about 4800 t; the purified residue does not have corrosive, leaching toxicity and other dangerous characteristics, and can be used for making bricks or building materials. Thermal phase separation technology is a comprehensive utilization and treatment technology with excellent engineering and environmental benefits, which has a high promotion value.

An artificial intelligence-based ecological index for prognostic evaluation of colorectal cancer.

BACKGROUND AND OBJECTIVE: In the tumor microenvironment (TME), the dynamic interaction between tumor cells and immune cells plays a critical role in predicting the prognosis of colorectal cancer. This study introduces a novel approach based on artificial intelligence (AI) and immunohistochemistry (IHC)-stained whole-slide images (WSIs) of colorectal cancer (CRC) patients to quantitatively assess the spatial associations between tumor cells and immune cells. To achieve this, we employ the Morisita-Horn ecological index (Mor-index), which allows for a comprehensive analysis of the spatial distribution patterns between tumor cells and immune cells within the TME. MATERIALS AND METHODS: In this study, we employed a combination of deep learning technology and traditional computer segmentation methods to accurately segment the tumor nuclei, immune nuclei, and stroma nuclei within the tumor regions of IHC-stained WSIs. The Mor-index was used to assess the spatial association between tumor cells and immune cells in TME of CRC patients by obtaining the results of cell nuclei segmentation. A discovery cohort (N = 432) and validation cohort (N = 137) were used to evaluate the prognostic value of the Mor-index for overall survival (OS). RESULTS: The efficacy of our method was demonstrated through experiments conducted on two datasets comprising a total of 569 patients. Compared to other studies, our method is not only superior to the QuPath tool but also produces better segmentation results with an accuracy of 0.85. Mor-index was quantified automatically by our method. Survival analysis indicated that the higher Mor-index correlated with better OS in the discovery cohorts (HR for high vs. low 0.49, 95% CI 0.27-0.77, P = 0.0014) and validation cohort (0.21, 0.10-0.46, < 0.0001). CONCLUSION: This study provided a novel AI-based approach to segmenting various nuclei in the TME. The Mor-index can reflect the immune status of CRC patients and is associated with favorable survival. Thus, Mor-index can potentially make a significant role in aiding clinical prognosis and decision-making.

Predictors of Return to Work After Thoracic Endovascular Aortic Repair in Patients With Type B Aortic Dissection.

OBJECTIVE: Type B aortic dissection (TBAD) is a life-threatening condition, and it takes heavy burden to family and society. Return to work (RTW) not only means patients' physical health but also demonstrates their mental well-being. Thoracic endovascular aortic repair (TEVAR) has been successful in treatment of TBAD patients. However, less studies have addressed on the social functional recovery of TBAD after TEVAR, especially for RTW. METHODS: From January 1, 2017 to January 1, 2021, TBAD patients who underwent TEVAR and completed a 12-month follow-up were retrospectively enrolled. Primary outcome was RTW. Patients' demographic, sociological, and clinical characteristics, and so on were recorded to analyze and demonstrate independent risk factors for RTW. RESULTS: Four hundred thirty-two TBAD patients (388 males) were enrolled with a mean age of 48.3±8.9 years (ranged from 19 to 60 years). The 12-month cumulative RTW rate was 62.7% (95% confidence interval [CI]: 57.2%-67.8%). Age <50 years (odds ratio [OR]=3.675, 95% CI: 1.436-9.405) was identified as independent protective factors for RTW, while preoperative job as manual workers (OR=0.101, 95% CI: 0.029-0.353), average annual income, <30 000 Chinese Yuan (CNY) [<4400 US dollar], (OR=0.186, 95% CI: 0.054-0.637), complicated TBAD (malperfusion) (OR=0.246, 95% CI: 0.092-0.659), and distal stent graft-induced new entry (SINE) (OR=0.218, 95% CI: 0.083-0.575, p=0.002) were identified as independent risk factors. CONCLUSION: Approximately 64% of our patients were able to RTW in the 12 months post-TEVAR for TBAD. Younger patients, patients with less physically demanding jobs, and patients with less complex surgeries were more likely to RTW. Based on these results, more can be done to facilitate the patient's ability and willingness to RTW after TEVAR. CLINICAL IMPACT: Type B aortic dissection (TBAD) is a life-threatening condition that poses significant burden on both individuals and society. The ability to return to work (RTW) not only reflects the patient's physical health but also indicates their mental well-being. Therefore, identifying risk factors for RTW and promoting the reintegration of TBAD patients into the workforce is crucial in clinical practice.To our knowledge, this study is the first to elucidate and predict the RTW outcomes of TBAD patients who underwent thoracic endovascular aortic repair (TEVAR).

Evaluation of the lower extremity blood supply in no-option critical limb ischemia patients with stem cell transplantation by time maximum intensity projection CT perfusion: A single-centre prospective study.

OBJECTIVES: Cell therapy has had satisfactory safety and efficacy outcomes for no-option critical limb ischaemia (NO-CLI) patients. In the current study, we aimed to compare the image quality of ischaemic lower limb blood vessels shown on volumetric CT-based time maximum intensity projection CT perfusion (t-MIP CTP) versus single-phase CTA (sCTA). We also tried to quantify the blood flow of the ischaemic lower extremity based on the t-MIP technique, not only to precisely show the dynamic change in blood flow from before to after cell therapy but also to detect any relationship between this change and patient prognosis. METHODS: A total of 31 patients with thromboangiitis obliterans (TAO)-induced NO-CLI who had been referred from the department of vascular surgery to undergo autologous stem cell transplantation into a single limb from January 2020 to March 2021 were prospectively enrolled in this study. Preoperative sCTA or t-MIP CTP and postoperative 1-month t-MIP CTP were performed in all patients. Clinical outcomes, including the 1-month ankle-brachial index (ABI) and 3-month CLI status, were also analysed. Image quality, including objective scores (attenuation, signal-to-noise ratio [SNR] and contrast-to-noise ratio [CNR]), subjective scores and collateral scores, was compared between preoperative sCTA and t-MIP CTP. Vascular volume was calculated as the total volume (mL) of lower limb arteries within the scanning range. All images and calculations were performed by 2 separate radiologists. Receiver operating characteristic curves were drawn to reveal the sensitivity and specificity of vascular volume and ABI in predicting prognosis. RESULTS: Both sCTA and t-MIP CTP images exhibited good quality for diagnosis. t-MIP CTP images showed significantly higher attenuation, SNR and CNR in all arterial segments (popliteal artery, anterior tibial artery, posterior tibial artery and peroneal artery). In subjective and collateral score evaluations, t-MIP CTP images were also significantly better than sCTA images (both p < .05). At 1 month after transplantation, both vascular volume and ABI showed significant improvement (both p < .01). At 3 months after transplantation, 38.71% of patients (12/31) achieved CLI relief (Rutherford class < 4). Through the receiver operating characteristic (ROC) curve, the 1-month vascular volume increase ratio showed better ability to predict the 3-month prognosis (radiologist 1: AUC, 0.757; sensitivity, 0.750; specificity, 0.840; radiologist 2: AUC, 0.803; sensitivity, 0.500; specificity, 1.000) than the 1-month ABI increase ratio (AUC, 0.607; sensitivity, 0.230; specificity, 0.820) or 1-month ABI (AUC, 0.410; sensitivity, 0.080; specificity, 0.580). CONCLUSION: t-MIP CTP showed significantly higher-quality images of ischaemic limb vascularity than sCTA. t-MIP CTP can reveal the anatomical information of collaterals more accurately, which is of great importance for NO-CLI patients undergoing cell transplantation. The 1-month vascular volume increase ratio can predict the 3-month prognosis more precisely on this basis.

Quantifying Urban Activities Using Nodal Seismometers in a Heterogeneous Urban Space.

Earth's surface is constantly vibrating due to natural processes inside and human activities on the surface of the Earth. These vibrations form the ambient seismic fields that are measured by sensitive seismometers. Compared with natural processes, anthropogenic vibrations dominate the seismic measurements at higher frequency bands, demonstrate clear temporal and cyclic variability, and are more heterogeneous in space. Consequently, urban ambient seismic fields are a rich information source for human activity monitoring. Improving from the conventional energy-based seismic spectral analysis, we utilize advanced signal processing techniques to extract the occurrence of specific urban activities, including motor vehicle counts and runner activities, from the high-frequency ambient seismic noise. We compare the seismic energy in different frequency bands with the extracted activity intensity at different locations within a one-kilometer radius and highlight the high-resolution information in the seismic data. Our results demonstrate the intense heterogeneity in a highly developed urban space. Different sectors of urban society serve different functions and respond differently when urban life is severely disturbed by the impact of the COVID-19 pandemic in 2020. The anonymity of seismic data enabled an unprecedented spatial and temporal resolution, which potentially could be utilized by government regulators and policymakers for dynamic monitoring and urban management.

Combination of Kaempferol and Docetaxel Induces Autophagy in Prostate Cancer Cells In Vitro and In Vivo.

Docetaxel is a first-line chemotherapy drug used to treat advanced prostate cancer, but patients who have used it often face the challenges of drug resistance and side effects. Kaempferol is a naturally occurring flavonol; our previous studies have confirmed that it has excellent anti-prostate activity. To investigate the anti-prostate cancer effects of docetaxel in combination with kaempferol, we conducted experiments at the cellular and whole-animal level. Plate cloning assays showed that the combination of docetaxel and kaempferol had a synergistic effect in inhibiting the proliferation of prostate cancer cells. The combination of these two compounds was found to induce autophagy in prostate cancer cells via transmission electron microscopy, and changes in the expression of autophagy-related proteins via Western blot assays also confirmed the occurrence of autophagy at the molecular level. We also confirmed the anti-prostate cancer effect of docetaxel in combination with kaempferol in vivo by establishing a mouse xenograft prostate cancer model. Autophagy-related proteins were also examined in mouse tumor tissues and verified the presence of autophagy in mouse tumor tissues. The above cellular and animal data suggest that docetaxel in combination with kaempferol has significant anti-prostate cancer effects and that it works by inducing autophagy in cells.

Detect the early-warning signals of diseases based on signaling pathway perturbations on a single sample.

BACKGROUND: During the pathogenesisof complex diseases, a sudden health deterioration will occur as results of the cumulative effect of various internal or external factors. The prediction of an early warning signal (pre-disease state) before such deterioration is very important in clinical practice, especially for a single sample. The single-sample landscape entropy (SLE) was proposed to tackle this issue. However, the PPI used in SLE was lack of definite biological meanings. Besides, the calculation of multiple correlations based on limited reference samples in SLE is time-consuming and suspect. RESULTS: Abnormal signals generally exert their effect through the static definite biological functions in signaling pathways across the development of diseases. Thus, it is a natural way to study the propagation of the early-warning signals based on the signaling pathways in the KEGG database. In this paper, we propose a signaling perturbation method named SSP, to study the early-warning signal in signaling pathways for single dynamic time-series data. Results in three real datasets including the influenza virus infection, lung adenocarcinoma, and acute lung injury show that the proposed SSP outperformed the SLE. Moreover, the early-warning signal can be detected by one important signaling pathway PI3K-Akt. CONCLUSIONS: These results all indicate that the static model in pathways could simplify the detection of the early-warning signals.

Kaempferol suppresses androgen-dependent and androgen-independent prostate cancer by regulating Ki67 expression.

PURPOSE: Kaempferol is a natural flavonoid that has been reported to be active against many cancers, including prostate cancer, breast cancer and colon cancer. In our previous study, we found kaempferol could act as a selective androgen receptor modulator, thereby suppress development of benign prostatic hyperplasia. This finding inspired us to further explore the effect and the mechanism of action of kaempferol on prostate cancer. METHODS: Plate clone formation assay was performed to detect the effect of kaempferol on cell proliferation. Flow cytometry was used to detect the impact of kaempferol on cell apoptosis and cell cycle distribution. qPCR, immunofluorescence staining, and enzyme-linked immunosorbent assay were used to detect the expression of gene and protein of Ki67 which is a biomarker of cell proliferation. RESULTS: In the present study, we found kaempferol could dramatically suppress androgen-dependent and androgen-independent prostate cancer cells proliferation and induce their apoptosis. Furthermore, we found that kaempferol induced cell cycle to be arrested at G1 phase in 22Rv1 cells but at S and G2 phase in PC-3 cells. In addition, we detected the mRNA and protein of Ki67 which is corresponding to the cell proliferation and found that kaempferol could significantly inhibit Ki67 expression at mRNA level but increase its expression at protein levels in both androgen-dependent and androgen-independent prostate cancer cells. CONCLUSION: Taken together, kaempferol inhibited the proliferation of androgen-dependent and androgen-independent prostate cancer cells by regulating the expression of Ki67. These findings further shed light on the mechanism of action of kaempferol on anti-prostate cancer.

Influences of Total Alkaloids in Caulis Mahoniae on Proliferation and Apoptosis of Cervical Cancer Cells and the Caspase-3 Expression.

The study aimed to investigate the influences of the active ingredient in Caulis Mahoniae, total alkaloids, on the proliferation and apoptosis of cervical cancer cells and the caspase-3 expression. The total alkaloids were extracted in vitro from Caulis Mahoniae, and cervical cancer HeLa cell lines were used as experimental objects. The half inhibitory concentration (IC50) of total alkaloids on HeLa cell lines was detected via the preliminary experiment, the influences of total alkaloids at different concentrations on the proliferation of HeLa cell lines were detected via methyl thiazolyl tetrazolium (MTT) assay, and the cell growth curve was plotted. Moreover, the cell cycle and apoptosis after treatment with total alkaloids at different concentrations were detected via flow cytometry, and the caspase-3 gene and protein expressions were detected via reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting. The IC50 of total alkaloids in Caulis Mahoniaeon HeLa cell lines was 12.5 µg/mL. With the gradual increase of concentration of total alkaloids in the treatment of cervical cancer cells, the inhibitory rate on cancer cells was gradually increased, and the proportion of cells in the G0/G1 phase was gradually decreased, while that in S and G2/M phases was gradually increased. Besides, with the increase in the concentration of total alkaloids, the apoptotic rate of cervical cancer cells was gradually increased, and both caspase-3 gene and protein expressions were also gradually increased. The total alkaloids extracted from Caulis Mahoniae can effectively inhibit the proliferation and promote the apoptosis of cervical cancer HeLa cells, which may be realized by promoting the expression of apoptosis-related factor caspase-3.

Total Flavonoids in Premna Fulva Craib Alleviates Brain Neurological Impairment and Influences Nrf2 and HO-1 Expressions in Rats with Ischemia-Reperfusion.

Total flavonoids in Premna fulva Craib (TFPFC) are a kind of flavonoid compound synthesized via photosynthesis extracted from Premna fulva Craib, which possess a strong anti-oxidative effect. Cerebral Ischemia-Reperfusion refers to the body's damage mainly caused by oxidative stress. This study aims to investigate the alleviating effect of TFPFC on brain neurological impairment and its influences on Nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase 1 (HO-1) expressions in rats with Ischemia-Reperfusion. The rat model of Ischemia-Reperfusion was established, and rats were treated with TFPFC or normal saline. At 24 h after reperfusion, the neurological score, volume of cerebral infarction and cerebral water content were analyzed in different groups. The influences of TFPFC treatment on the proliferative activity and apoptosis of oxygen and glucose deprivation/reoxygenation (OGD/R) neural stem cells were detected via methyl thiazolyl tetrazolium (MTT) assay and flow cytometry. Moreover, the malondialdehyde (MDA) level and superoxide dismutase (SOD) activity were measured to evaluate the oxidative stress effect. The influences of TFPFC treatment on the protein and messenger ribonucleic acid (mRNA) expressions of Nrf2 and HO-1 were analyzed using reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting. The TFPFC treatment alleviated the neurological impairment in rats after Ischemia-Reperfusion and reduced the volume of cerebral infarction and cerebral edema status in rats with Ischemia-Reperfusion. TFPFC increased the proliferative activity of OGD/R neural stem cells and decreased damage and apoptosis. In addition, the TFPFC treatment reduced the MDA level, improved the SOD activity, and up-regulated the protein and mRNA expressions of Nrf2 and HO-1. The TFPFC treatment may improve oxidative damage and protect the nervous system through the up-regulation of expressions of transcription factors Nrf2 and HO-1.