Puerarin-V Improve Mitochondrial Respiration and Cardiac Function in a Rat Model of Diabetic Cardiomyopathy via Inhibiting Pyroptosis Pathway through P2X7 Receptors.

There is a new form of puerarin, puerarin-V, that has recently been developed, and it is unclear whether puerarin-V has a cardioprotective effect on diabetic cardiomyopathy (DCM). Here, we determined whether puerarin-V had any beneficial influence on the pathophysiology of DCM and explored its possible mechanisms. By injecting 30 mg/kg of STZ intraperitoneally, diabetes was induced in rats. After a week of stability, the rats were injected subcutaneously with ISO (5 mg/kg). We randomly assigned the rats to eight groups: (1) control; (2) model; (3) metformin; (4-6) puerarin-V at different doses; (7) puerarin (API); (8) puerarin injection. DCM rats were found to have severe cardiac insufficiency (arrythmia, decreased LVdP/dt, and increased E/A ratio). In addition, cardiac injury biomarkers (cTn-T, NT-proBNP, AST, LDH, and CK-MB), inflammatory cytokines (IL-1ß, IL-18, IL-6, and TNF-α), and oxidative damage markers (MDA, SOD and GSH) were markedly increased. Treatment with puerarin-V positively adjusts these parameters mentioned above by improving cardiac function and mitochondrial respiration, suppressing myocardial inflammation, and maintaining the structural integrity of the cardiac muscle. Moreover, treatment with puerarin-V inhibits the P2X7 receptor-mediated pyroptosis pathway that was upregulated in diabetic hearts. Given these results, the current study lends credence to the idea that puerarin-V can reduce myocardial damage in DCM rats. Furthermore, it was found that the effect of puerarin-V in diabetic cardiomyopathy is better than the API, the puerarin injection, and metformin. Collectively, our research provides a new therapeutic option for the treatment of DCM in clinic.

3q26.2/EVI1 rearrangement is associated with poor prognosis in classical Philadelphia chromosome-negative myeloproliferative neoplasms.

Classical Philadelphia chromosome-negative myeloproliferative neoplasms are a group of closely related myeloid disorders with different histologic features and clinical presentations at an early stage, but all later develop into a similar fibrotic stage with variable risk of acute transformation. The significance of 3q26.2/EVI1 rearrangement has been well recognized in acute myeloid leukemia, myelodysplastic syndrome, and chronic myeloid leukemia. However, the clinical importance of 3q26.2/EVI1 rearrangement in classical Philadelphia chromosome-negative myeloproliferative neoplasms is unknown. Here we reported 15 patients with classical Philadelphia chromosome-negative myeloproliferative neoplasms showing 3q26.2 rearrangement, including inv(3)(q21q26.2) (n=6), t(3;21)(q26.2;q22)(n=4), t(3;3)(q21;q26.2)(n=3), inv(3)(q13.3q26.2)(n=1), and t(3;12)(q26.2;p13)(n=1). In addition to 3q26.2 rearrangement, 9 of 15 cases had other concurrent karyotypical abnormalities, including -7/7q- and -5/5q-. There were 8 men and 7 women with a median age of 59 years (range, 35-79 years) at initial diagnosis of myeloproliferative neoplasms: 8 patients had primary myelofibrosis, 4 had polycythemia vera, and 3 had essential thrombocythemia. JAK2 V617F mutation was detected in 8/14 patients, including 4/4 with polycythemia vera. The median interval from the initial diagnosis of myeloproliferative neoplasms to the detection of 3q26.2 rearrangement was 44 months (range, 1-219 months). At time of emergence of 3q26.2 rearrangement, 11 patients were in blast phase and 2 patients had increased blasts (6-19%). Dyspoiesis, predominantly in megakaryocytes, were detected in all patients with adequate specimens at time of 3q26.2 rearrangement. Following 3q26.2 rearrangement, 12 patients received chemotherapy, but none of them achieved complete remission. Of 14 patients with follow-up information, all died with a median overall survival time of only 3 months (range 0-14 months) after the emergence of 3q26.2 rearrangement. In summary, 3q26.2 rearrangement in classical Philadelphia chromosome-negative myeloproliferative neoplasms is associated with other concurrent cytogenetic abnormalities, a rapid disease progression and blast transformation, a poor response to chemotherapy and a dismal prognosis.

Clinical implications of cytogenetic heterogeneity in multiple myeloma patients with TP53 deletion.

TP53 deletion (ΔTP53) in myeloma is known to be a high-risk finding associated with poorer prognosis. The prognostic impact of underlying cytogenetic heterogeneity in patients with myeloma associated with ΔTP53 is unknown. We studied 90 patients with myeloma associated with ΔTP53 identified by interphase fluorescence in situ hybridization and assessed the impact of karyotype and coexisting alterations of IGH, RB1, and CKS1B. There were 54 men and 36 women with a median age of 59 years (range 38-84); 14 patients had a normal karyotype (NK/ΔTP53), 73 had a complex karyotype (CK/ΔTP53), and 3 had a non-complex abnormal karyotype. Patients with CK/ΔTP53 showed a significantly poorer overall survival compared with patients with NK/ΔTP53 (P=0.0243). Furthermore, in the CK/ΔTP53 group, patients with IGH rearrangement other than t(11;14)(q13;q32)/CCND1-IGH, designated as adverse-IGH, had an even worse outcome (P=0.0045). In contrast, RB1 deletion, CKS1B gain, ploidy, additional chromosome 17 abnormalities, or ΔTP53 clone size did not impact prognosis. Stem cell transplant did not improve overall survival in either the NK/ΔTP53 or CK/ΔTP53 (P=0.8810 and P=0.1006) groups, but tandem stem cell transplant did improve the overall survival of patients with CK/ΔTP53 (P=0.0067). Multivariate analysis confirmed in this cohort that complex karyotype (hazard ratio 1.976, 95% CI 1.022-3.821, P=0.043), adverse-IGH (hazard ratio 3.126, 95% CI 1.192-8.196, P=0.020), and tandem stem cell transplant independently correlate with overall survival (hazard ratio 0.281, 95% CI 0.091-0.866, P=0.027). We conclude that comprehensive genetic assessment adds to TP53 status in the risk stratification of myeloma patients.

Additional-structural-chromosomal aberrations are associated with inferior clinical outcome in patients with hyperdiploid multiple myeloma: a single-institution experience.

Multiple myeloma is cytogenetically heterogeneous and a hyperdiploid karyotype is considered currently to have standard risk. In this study, we investigated the clinical impact of additional-structural-chromosomal aberrations assessed by chromosome analysis in 284 patients with a hyperdiploid karyotype that were subdivided into four groups based on the complexity of additional-structural-chromosomal aberrations: group 1, no additional-structural-chromosomal aberrations (n=35); group 2, one additional-structural-chromosomal aberration (n=46); group 3, two additional-structural-chromosomal aberrations (n=39); group 4, ≥three additional-structural-chromosomal aberrations (n=164). Clinicopathological data among these groups showed no differences, except patients in group 1 had higher hemoglobin (P=0.031) and albumin (P=0.045) levels. The median follow-up was 55 months (range, 3-221). The median overall survival of patients in groups 1-4 was negatively correlated with the number of the additional-structural-chromosomal aberrations: 98, 76, 61, and 48 months, respectively (P<0.0001). In group 4, CKS1B gain, RB1, or TP53 deletions had no additional impact on overall survival; however, trisomy 3 or 15 conferred a much better overall survival, and monosomy 13 and 14 predicted a worse outcome. In addition, the overall survival of patients in groups 3 and 4 was similar to a subset of high-risk multiple myeloma cases (n=21) (P=0.387). About 192 (67.6%) patients who received stem cell transplantation did not show improved overall survival compared with non-stem cell transplantation patients (n=92; P=0.142) overall; however, they did show significantly improved overall survival in patients with refractory disease in group 4 (P=0.0084). Multivariate analysis showed that two or more additional-structural-chromosomal aberrations (P<0.0001), stages (P=0.02 and P=0.002) and relapsed disease (P=0.009) negatively impacted the overall survival. We conclude that hyperdiploid karyotypes in multiple myeloma are associated with additional-structural-chromosomal aberrations and a greater number of additional-structural-chromosomal aberrations predicts poorer clinical outcome. A hyperdiploid karyotype with ≥2 additional-structural-chromosomal aberrations at chromosomal level should be considered an independent high-risk factor.

Prognostic significance of cytogenetic abnormalities in T-cell prolymphocytic leukemia.

T-cell prolymphocytic leukemia (T-PLL) is an aggressive mature T-cell neoplasm. The most common cytogenetic abnormality associated with T-PLL is inv(14)(q11.2q32) involving TCL1, but other abnormalities also have been reported. In this study, we correlated cytogenetic abnormalities with clinical outcome in 97 T-PLL patients, including 66 men and 31 women with a median age of 63 years (range, 34-81). Twenty-seven patients had a normal karyotype (NK), one had two chromosomal aberrations, and 69 had a complex karyotype (CK). Patients with a CK had poorer overall survival (OS) than patients with a NK (P = .0016). In the CK group, the most common aberrations involved 14q (n = 45) and 8q (n = 38). Additional deletions of chromosomes 17p, 11q, 6q, 12p, 13q were observed frequently. No individual cytogenetic abnormality impacted OS. Patients with ≥5 aberrations had an OS of 11 months versus 22 months in patients with <5 aberrations (P = 0.0132). Fluorescence in situ hybridization for TCL1 successfully performed in 27 cases showed rearrangement in 8/10 (80%) NK versus 16/17 (94%) CK cases. OS of patients with TCL1 rearrangement and/or 14q aberrations was not significantly different from patients without TCL1 rearrangement and 14q aberrations (P = .3467). Patients with refractory disease showed worse OS in both the NK and CK groups (P = .0014 and P < .0001, respectively), compared with patients who achieved remission but then relapsed. Stem cell transplantation did not appear to improve OS regardless of karyotype complexity. In conclusion, patients with T-PLL often have a CK which is a poor prognostic factor, particularly in patients with ≥5 cytogenetic aberrations.

Aberrant Factors of Fibrinolysis and Coagulation in Pancreatic Cancer.

Aberrant factors associated with fibrinolysis and thrombosis are found in many cancer patients, which can promote metastasis and are associated with poor prognosis. The relationship between tumor-associated fibrinolysis and thrombosis is poorly understood in pancreatic cancer. This review provides a brief highlight of existing studies that the fibrinolysis and coagulation systems were activated in pancreatic cancer patients, along with aberrant high concentrations of tissue plasminogen activator (t-PA), urine plasminogen activator (u-PA), D-dimer, fibrinogen, or platelets. These factors cooperate with each other, propelling tumor cell shedding, localization, adhesion to distant metastasis. The relationship between thrombosis or fibrinolysis and cancer immune escape is also investigated. In addition, the potential prevention and therapy strategies of pancreatic cancer targeting factors in fibrinolysis and coagulation systems are also been discussed, in which we highlight two effective agents aspirin and low-molecular weight heparin (LMWH). Summarily, this review provides new directions for the research and treatment of pancreatic cancer.

Clinical implications of cytogenetic heterogeneity in Philadelphia chromosome positive (Ph+) adult B cell acute lymphoblastic leukemia following tyrosine kinase inhibitors and chemotherapy regimens.

We retrospectively studied a cohort of 144 adults with Philadelphia chromosome/BCR-ABL1 positive B acute lymphoblastic leukemia (Ph + B-ALL) to assess the clinical implications of cytogenetic heterogeneity in this disease. The study group included 85 men and 59 women that were sorted into 6 subgroups based on karyotypic findings in the stemline as follows: 32 patients with t(9;22) as a sole aberration, 23 with t(9;22) plus 1 additional chromosomal abnormality (ACA), 26 with t(9;22) as part of a complex karyotype, 18 showing a variant-/complex- t(9;22), 30 with t(9;22) as the stemline with ACAs in the sideline(s), and 15 patients who had the t(9;22) and hyperdiploidy. In 89 patients 1 clone was identified; 41 had 2 clones and 14 had ≥ 3 clone(s). The median overall survival (OS) was 25.6 months and the median relapse-free survival (RFS) was 20.6 months. Patients with variant-/complex- t(9;22) had poorer OS and RFS when compared with all other subgroups combined (P = 0.0018 and P = 0.0049, respectively). In addition, patients with ≥ 2 clones had worse OS and RFS than patients with 1 clone (P = 0.0179 and P = 0.0429, respectively). Multivariate analysis confirmed that variant-/complex-t(9;22) and clone number are independent risk factors. We suggest that conventional chromosomal analysis is of clinical importance for risk stratification of B-ALL patients.

MET amplification assessed using optimized FISH reporting criteria predicts early distant metastasis in patients with non-small cell lung cancer.

To investigate the prognostic impact of MET copy number (MET-CN) in patients with non-small cell lung cancer (NSCLC), we retrospectively reviewed clinical and pathologic data of NSCLC patients whose tumors were assessed for MET-CN using fluorescence in situ hybridization (FISH). We correlated MET-CN status with patient overall survival (OS) and optimized MET-FISH reporting criteria. The study group included 384 patients with NSCLC of which 88% were adenocarcinoma and 55.7% of patients had distant metastases. There were 170 patients with stages I-III and 214 patients with stage IV disease. Based on the MET-CN and MET/CEP7 ratio the patients were classified into 3 categories: MET-amplification (METamp): MET/CEP7 ≥ 2 or MET-CN ≥ 5; MET-CN-gain (METcng): MET-CN ≥ 4 to < 5; and MET-negative (METneg): MET-CN < 4. METamp was associated with high fatality (P=.036) and stage IV tumors (P=.038). In patients with stages I-III NSCLC, patients in the METamp category had the shortest OS (P=.015) and more often developed distant metastases within 1 year (P=.004). In patients with stage IV tumors, METamp did not further impact the OS. Patients in the METcng category had the longest OS (P=.053). Multivariate analysis confirmed METamp to be an independent high-risk factor (HR 3.26; P=.026) and predicted earlier progression to distant metastasis (HR 4.86; P=.001). In conclusion, we suggest that the MET-FISH criteria presented optimizes risk stratification by defining 3 categories of NSCLC patients. METamp is an independent risk factor predicting early distant metastasis and patients with METcng could represent a lower-risk group.

Myeloproliferative neoplasms with t(8;22)(p11.2;q11.2)/BCR-FGFR1: a meta-analysis of 20 cases shows cytogenetic progression with B-lymphoid blast phase.

Rearrangements of FGFR1 result in the 8p11 myeloproliferative syndrome, a group of rare diseases that features a myeloproliferative neoplasm (MPN) that commonly progresses to lymphoblastic leukemia/lymphoma or acute myeloid leukemia. The most common partner of FGFR1 is ZMYM2, and patients with the ZMYM2-FGFR1 fusion often present with MPN and T-lymphoblastic lymphoma. There are 14 other partners that can fuse with FGFR1, and of interest is the BCR-FGFR1 fusion that results from t(8;22)(p11.2;q11.2). Patients with t(8;22) often show leukocytosis and present with an MPN resembling chronic myeloid leukemia or very rarely, with B-lymphoblastic leukemia (B-ALL). In this study, we analyzed the clinicopathological, cytogenetic, and molecular features of 2 new patients with the t(8;22)(p11.2;q11.2)/BCR-FGFR1 who presented with B-ALL. An underlying MPN became apparent when a morphologic remission of B-ALL was achieved after chemotherapy. We subsequently reviewed the literature and identified 18 additional cases reported with B-ALL in a background MPN or with the MPN as a chronic phase. Our data suggest that the t(8;22)(p11.2;q11.2)/BCR-FGFR1 may arise from a myeloid/B progenitor cell. It is important to recognize that neoplasms carrying the t(8;22)/BCR-FGFR1, although rare, can commonly with B lymphoblastic leukemia at the initial diagnosis, which could distract one from recognizing a possible underlying 8p11 myeloproliferative syndrome.

Solid-phase microextraction-gas chromatographic-mass spectrometric analysis of volatile compounds from Curcuma wenyujin Y.H. Chen et C. Ling.

A solid-phase microextraction coupled with gas chromatography-mass spectrometry (SPME-GC-MS) for analysis of the volatile compounds from Curcuma wenyujin Y.H. Chen et C. Ling is described. SPME parameters (fiber type, extraction temperature and time, headspace volume and desorption time) and GC conditions were tested. The powdered sample of C. wenyujin Y.H. Chen et C. Ling was directly analyzed by SPME-GC-MS and 72 compounds were identified. The results from SPME-GC-MS were compared with those obtained from steam distillation gas chromatography-mass spectrometry (SD-GC-MS) with a good agreement. The results show that SPME-GC-MS method is a fast, simple and efficient way for the analysis of volatile components from traditional Chinese medicines (TCMs).

Headspace solvent microextraction-gas chromatography-mass spectrometry for the analysis of volatile compounds from Foeniculum vulgare Mill.

A novel and rapid headspace solvent microextraction followed by gas chromatography-mass spectrometry (HSME-GC-MS) for the analysis of the volatile compounds of Foeniculum vulgare Mill is described. HSME parameters including extracting solvent, extraction temperature and time, headspace volume and particle size were optimized. As a result, benzyl alcohol was finally used for the extraction at 70 degrees C for 20 min with headspace volume of 12.1 ml and particle size of 120 mesh. Under the determined conditions, the powered samples of Foeniculum vulgare Mill were directly applied for the analysis. A comparison of HSME-GC-MS, solid phase microextraction (SPME)-GC-MS and steam distillation (SD)-GC-MS methods was made and showed that the HSME-GC-MS method was simple, inexpensive and effective and can be used for the analysis of volatile compounds in traditional Chinese medicines (TCMs).

Development of multiple malignancies following long-term glucocorticoid therapy in a patient with leukocytoclastic vasculitis: A case report.

Leukocytoclastic vasculitis (LCV) is a neutrophilic inflammation of the blood vessels. LCV may present as a paraneoplastic syndrome occurring before, synchronously with, or after the diagnosis of malignancy. In this study, we report a unique case of multiple malignancies developing simultaneously in a patient with a long history of LCV. The patient was originally diagnosed with LCV and received long-term glucocorticoid treatment. After 11 years of therapy, the patient developed three primary malignancies, including small-cell lung carcinoma, gastric adenocarcinoma and colonic adenocarcinoma. It is likely that LCV was not a paraneoplastic syndrome in this case, but rather an independent process, and the development of multiple cancers is likely associated with the long-term glucocorticoid treatment, which caused imbalance of the immune system. Although the development of cancer during the course of glucocorticoid treatment is very rare, clinicians must be aware of this possible association and immunodysregulation may play a role in this context.

Tounong Powder () extracts induce G1 cell cycle arrest and apoptosis in LoVo cells.

OBJECTIVE: To further explore the anti-cancer effect of Tounong Powder () extracts (TNSEs) on human colon cancer LoVo cells and examine the possible molecular mechanisms. METHODS: The contents of TNSEs were determined by liquid chromatograph-mass spectrometer (LC-MS) analysis after extraction with water and methanol. Variations of cell morphological features were observed using fluorescence microscopy. Cytotoxicity was determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell cycle distribution and apoptosis were analyzed using flow cytometry at different TNSE doses (0, 62.5, 125, or 250 µg/mL). Protein expressions of phosphatidylinositol 3-kinase (PI3K), phosphate protein kinase B (p-AKT), phosphate mammalian target of rapamycin (p-mTOR), p-p70s6k1, cleaved caspase-9 and -3 were detected using Western blot analysis. RESULTS: TNSEs induced cell growth inhibition in a concentration- and time-dependent manner. Flow cytometric analysis showed apoptotic cells and cell cycle arrest at the G phase after TNSEs treatment compared with controls. Furthermore, TNSEs significantly down-regulated the proteins PI3K, p-AKT, p-mTOR, and p-p70s6k1, and up-regulated the proteins cleaved caspase-9 and -3 dosedependently, as determined by Western blot. CONCLUSIONS: TNSEs reduced LoVo cell proliferation, and caused apoptosis and cell-cycle arrest in LoVo cells. This effect might be associated with regulation of the PI3K/AKT signaling pathway.

Coexistence of iAMP21 and ETV6-RUNX1 fusion in an adolescent with B cell acute lymphoblastic leukemia: literature review of six additional cases.

BACKGROUND: Intrachromosomal amplification of chromosome 21 (iAMP21) results from breakage-fusion-bridge cycles and chromothripsis is a distinct marker of a subgroup of B cell acute lymphoblastic leukemia (B-ALL) cases associated with a poor prognosis. iAMP21 accounts for 2% of pediatric B-ALL and occurs predominantly in older children or adolescents. ETV6-RUNX1 fusion, resulting from t(12;21)(p13;q22), is associated with an excellent outcome in younger children with B-ALL. Coexistence of iAMP21 with ETV6-RUNX1 fusion is extremely rare with limited clinical information available. RESULTS: We report the case of an 18-year old Caucasian man diagnosed with ETV6-RUNX1 fusion positive B-ALL. He was treated with intensive chemotherapy and achieved remission for 6 months before relapse, 15 months after the initial diagnosis. G-band karyotyping and Fluorescence in situ hybridization (FISH) analyses performed on bone marrow revealed complex abnormalities: 41,X,-Y,der(3)t(3;20)(p11.2;q11.2),-4,t(5;22)(q32;q11.2),del(9)(p13),dic(9;17)(p13;p11.2),t(12;21)(p13;q22),der(14)t(14;17)(p11.2;q11.2),der(17;22)(q11.2;q11.2),-20,add(21)(q22),-22[4]/46,XY[15] with an iAMP21 and an ETV6-RUNX1. Additional molecular studies confirmed ETV6-RUNX1 fusion and with a TP53 mutation. High-resolution single nucleotide polymorphism microarray (SNP array) revealed the iAMP21 to be chromothripsis of 21q and subsequent metaphase FISH further delineated complex genomic aberrations. Although the patient received intensive chemotherapy with allogenic stem cell transplant, he died 26 months after initial diagnosis. We searched the literature and identified six cases showing coexisting iAMP21 and ETV6-RUNX1. The median age for these six patients was 10 years (range, 2-18) and males predominated. The median overall survival (OS) was 28 months. CONCLUSIONS: Patients with B-ALL associated with both iAMP21 and ETV6-RUNX1 tend to be older children or adolescents and have a poor prognosis.

Autophagy Protects from Raddeanin A-Induced Apoptosis in SGC-7901 Human Gastric Cancer Cells.

Raddeanin A (RA) is an extractive from Anemone raddeana Regel, a traditional Chinese medicine. The aim of this study is to assess the efficacy of RA against human gastric cancer (GC) cells (SGC-7901) and explore its mechanism. MTT assay showed that RA inhibition of proliferation of SGC-7901 cells increased in a dose-dependent manner. Flow cytometry analysis and Hoechst 33258 staining showed that RA induced apoptosis on SGC-7901 cells. Meanwhile, it induced autophagy. Western blotting analysis showed that the RA induces apoptosis and autophagy by activating p38 MAPK pathway and inhibiting mTOR pathway. Further studies showed that autophagy inhibition could protect from RA-induced apoptosis in SGC-7901 cells. In conclusion, RA can induce SGC-7901 cell apoptosis and autophagy by activating p38 MAPK pathway. And autophagy can protect SGC-7901 cells from apoptosis induced by RA.

The effect of tou nong san on transplanted tumor growth in nude mice.

Tou Nong San (TNS) is a traditional Chinese medicinal decoction used to treat sores and carbuncles. It contains four herbal drugs and one animal medicine: Radix Astragaliseu Seu Hedysari, Angelica sinensis, Ligustici Chuanxiong, Spina Gleditsiae, and stir-baked Squama Manis. Previous studies have shown that it has anticancer effects. This report validates in vivo antitumor properties of TNS. The compounds contained in TNSE were confirmed by liquid chromatographmass spectrometer (LC-MS) analysis. The in vivo antitumor activity of TNS extract (TNSE) was tested by feeding it to athymic mice harboring a human colonic tumor subcutaneous xenograft. Toxicity was monitored by recording behavior and weight parameters. Seven compounds were detected in TNSE by LC-MS. TNSE was fed to athymic mice for 2 weeks. No adverse reactions were reported. Compared to the control group, administration of TNSE to tumor bearing mice significantly reduced both tumor weight and volume. The expressions of p-PI3K, p-AKT, p-mTOR, p-p70s6k1, VEGF, and CD31 were significantly reduced, the expression levels of cleaved Caspase-9 and cleaved Caspase-3 were significantly increased in the TNSE groups compared to the control group as determined by western blot and immunohistochemistry. TNSE produced anticolonic cancer effects and the underlying mechanisms involved inhibition of the PI3K/AKT signal transduction pathway, inhibition of angiogenesis, and promotion of apoptotic proteins.

Tounongsan extract induces apoptosis in cultured Raji cells.

OBJECTIVE: To investigate the effects of Tounongsan () extract (TNSE) on proliferation and apoptosis of the human lymphoma cell line Raji and its possible mechanism of action. METHODS: The viability of TNSE-treated Raji cells was measured by a 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Cell apoptosis was determined by flow cytometry. The molecular mechanisms of TNSE-mediated apoptosis were further investigated by reverse transcription-polymerase chain reaction (RT-PCR) analysis of the mRNA expression of nuclear factor κB (NF-κB), Bcl-xL, Bcl-2-associated death promoter (Bad), caspase-9 and caspase-3. Western blotting was used to detect the protein expressions of NF-κB, Bad, cleaved caspase-9 and cleaved caspase-3. RESULTS: TNSE inhibited Raji cell proliferation in dose- and time-dependent manners. After 48-h treatment with various concentrations of TNSE (125, 250 and 500 µg/mL), the apoptosis rates of Raji cell were 12.23%±1.98% (P<0.05), 20.97%±3.96% (P<0.01) and 30.4%±4.87% (P<0.01), respectively, compared with those of the control (6.02%±1.01%). RT-PCR demonstrated that NF-κB mRNA expression was significantly downregulated in Raji cells treated with 250 µg/mL TNSE for 48 h (P<0.05), while Bad, caspase-9 and caspase-3 mRNA levels were upregulated (P<0.05). Moreover, TNSE treatment resulted in downregulation of NF-κB protein expression and strikingly upregulated protein expressions of Bad, cleaved caspase-9, cleaved caspase-3 in a dose-dependent manner, as determined by Western blot. CONCLUSION: TNSE exhibits significant anti-proliferative and apoptotic effects in Raji cells, which may be involved in regulation of NF-κB and Bad, and activation of caspase-9 and caspase-3.

Synergistic effect of a combination of nanoparticulate Fe3O4 and gambogic acid on phosphatidylinositol 3-kinase/Akt/Bad pathway of LOVO cells.

BACKGROUND: The present study evaluated whether magnetic nanoparticles containing Fe(3)O(4) could enhance the activity of gambogic acid in human colon cancer cells, and explored the potential mechanisms involved. METHODS: Cytotoxicity was evaluated by MTT assay. The percentage of cells undergoing apoptosis was analyzed by flow cytometry, and cell morphology was observed under both an optical microscope and a fluorescence microscope. Reverse transcriptase polymerase chain reaction and Western blot assay were performed to determine the transcription of genes and expression of proteins, respectively. RESULTS: Gambogic acid could inhibit proliferation of LOVO cells in a dose-dependent and time-dependent manner and induce apoptosis, which was dramatically enhanced by magnetic nanoparticles containing Fe(3)O(4). The typical morphological features of apoptosis in LOVO cells were observed after treatment comprising gambogic acid with and without magnetic nanoparticles containing Fe(3)O(4). Transcription of cytochrome c, caspase 9, and caspase 3 genes was higher in the group treated with magnetic nanoparticles containing Fe(3)O(4) and gambogic acid than in the groups that received gambogic acid or magnetic nanoparticles containing Fe(3)O(4), but transcription of phosphatidylinositol 3-kinase, Akt, and Bad genes decreased. Notably, expression of cytochrome c, caspase 9, and caspase 3 proteins in the group treated with gambogic acid and magnetic nanoparticles containing Fe(3)O(4) was higher than in the groups receiving magnetic nanoparticles containing Fe(3)O(4) or gambogic acid, while expression of p-PI3K, p-Akt, p-Bad, pro-caspase 9, and pro-caspase 3 degraded. CONCLUSION: Magnetic nanoparticles containing Fe(3)O(4) can enhance apoptosis induced by gambogic acid which may be closely related to regulation of the PI3K/Akt/Bad pathway in the treatment of human colon cancer.