Enhanced anticancer activity and endocytic mechanisms by polymeric nanocarriers of n-butylidenephthalide in leukemia cells

Purpose The purpose of this study was to investigate the antitumor mechanisms of n-butylidenephthalide (BP) and to further examine the delivery efficacy of polycationic liposome containing PEI and polyethylene glycol complex (LPPC)-encapsulated BP in leukemia cells. Method MTS, flow cytometric and TUNEL assays were performed to assess cell viability and apoptosis. BP and BP/LPPC complex delivery efficiency was analyzed by full-wavelength fluorescent scanner and fluorescence microscope. The expressions of cell cycle- and apoptosis-related proteins were conducted by Western blotting. Results The results showed that BP inhibited leukemia cell growth by inducing cell cycle arrest and cell apoptosis. LPPC-encapsulated BP rapidly induced endocytic pathway activation, resulting in the internalization of BP into leukemia cells, causing cell apoptosis within 1 h. Conclusions LPPC encapsulation enhanced the cytotoxic activity of BP and did not influence the effects of BP induction that suggested LPPC-encapsulated BP might be developed as anti-leukemia drugs in future (AU)

Enhanced anticancer activity and endocytic mechanisms by polymeric nanocarriers of n-butylidenephthalide in leukemia cells.

PURPOSE: The purpose of this study was to investigate the antitumor mechanisms of n-butylidenephthalide (BP) and to further examine the delivery efficacy of polycationic liposome containing PEI and polyethylene glycol complex (LPPC)-encapsulated BP in leukemia cells. METHODS: MTS, flow cytometric and TUNEL assays were performed to assess cell viability and apoptosis. BP and BP/LPPC complex delivery efficiency was analyzed by full-wavelength fluorescent scanner and fluorescence microscope. The expressions of cell cycle- and apoptosis-related proteins were conducted by Western blotting. RESULTS: The results showed that BP inhibited leukemia cell growth by inducing cell cycle arrest and cell apoptosis. LPPC-encapsulated BP rapidly induced endocytic pathway activation, resulting in the internalization of BP into leukemia cells, causing cell apoptosis within 1 h. CONCLUSIONS: LPPC encapsulation enhanced the cytotoxic activity of BP and did not influence the effects of BP induction that suggested LPPC-encapsulated BP might be developed as anti-leukemia drugs in future.

Telomerase activity in human hepatocellular carcinoma: parallel correlation with human telomerase reverse transcriptase (hTERT) mRNA isoform expression but not with cell cycle modulators or c-Myc expression.

BACKGROUND: To explore the possible regulatory mechanisms of telomerase, we examined the telomerase activity (TA), expression of human telomerase RNA (hTR), human telomerase reverse transcriptase (hTERT) mRNA isoforms and cell cycle modulators in human hepatocellular carcinoma (HCC) cell lines (J5, J7) and a normal human immortalized hepatic epithelial cell line (Chang-liver). METHODS: The cell lines were chemically synchronized in either G1, G1/S, G2/M or M phases. TA was measured by polymerase chain reaction (PCR)-based telomerase repeat amplification protocol assay. The hTR and hTERT mRNA levels were analyzed by reverse transcriptase-polymerase chain reaction. Western blotting and immunocytochemistry were used to assay the cell cycle modulators. RESULTS: The TA of J5, J7 and Chang-liver cell lines tested was highest in M phase. The expression level of hTERT mRNA associated with the highest TA detected in the M phase of HCC cell lines. Chang-liver expressed markedly less TA and hTERT mRNA than J5 or J7. The elevated TA and expression of hTERT mRNA isoforms in M phase of HCC cell lines did not significantly correlate with that of the cell cycle modulators and c-Myc. CONCLUSIONS: The results implicate that regulation of TA is related to hTERT mRNA isoform expression, and that regulation is different between the cell immortalization and tumorigenesis.

Polymorphism of the beta(2)-adrenoceptor in COPD in Chinese subjects.

STUDY OBJECTIVES: To assess the frequencies of three polymorphisms at amino acid positions 16, 27, and 164 of the beta(2)-adrenoceptor (beta(2)-AR) gene and their effects on COPD patients. DESIGN: Prospective, case-control study PATIENTS: Sixty-five patients with COPD and 41 healthy subjects were included. MEASUREMENTS: Polymorphisms of the beta(2)-AR coding block were delineated using an allele-specific polymerase chain reaction (PCR) approach. The allele-specific PCR technique was verified by direct dideoxy sequencing of PCR products. Pulmonary function tests were performed in all patients. RESULTS: The Arg16 beta(2)-AR polymorphism was less prevalent in COPD patients than in healthy populations (p = 0.01). A significant correlation (p < 0.018) between the Gln27 beta(2)-AR polymorphism and FEV(1) percent predicted value was found. Patients with the Gln27 polymorphism had a higher percentage of low FEV(1) percent predicted than did patients with the GlnGlu and GluGlu variants. CONCLUSIONS: The polymorphism of Gly16 may increase the patient's susceptibility to the development of COPD. The Gln27 beta(2)-AR polymorphism may be associated with the severity of COPD in a Chinese population.

Sensitive measurement of polyethylene glycol-modified proteins.

An IgM monoclonal antibody (AGP3) against polyethylene glycol (PEG) was used to assay PEG-modified proteins by ELISA. PEG-modified beta-glucuronidase could be measured at concentrations as low as 15 ng/mL, corresponding to 750 pg (1.8 fmol) of conjugate. This ELISA should be generally applicable to all PEG-modified proteins because AGP3 binds the backbone of the PEG chain independent of the linker used for PEG attachment.

Microsatellite alterations on human chromosome 11 in in situ and invasive breast cancer: a microdissection microsatellite analysis and correlation with p53, ER (estrogen receptor), and PR (progesterone receptor) protein immunoreactivity.

BACKGROUND AND OBJECTIVES: Microsatellite instability (MSI) has been documented in a subset of sporadic tumors. Loss of heterozygosity (LOH) on chromosome 11 loci in breast cancer is a frequent event. The purpose of the present study is to examine the incidence of microsatellite alterations in in situ and invasive human breast carcinoma and to clarify their significance in regulating the dynamics of cancer progression. METHODS: Four highly polymorphic (CA)n repeat microsatellites were used to determine microsatellite alterations in ten ductal carcinoma in situ (DCIS) and 19 invasive ductal carcinoma (IDC). To investigate the expression of p53, ER (estrogen receptor), and PR (progesterone receptor) association with MSI, immunohistochemistry staining was applied. RESULTS: MSI were detected in 20% (2/10) of DCIS and in 47.4% (9/19) of IDC. The frequency of MSI in IDC was significantly higher than that in DCIS (P < 0.001). Also, the MSI seemed to correlate with clinical stage (P = 0.0001) and tumor size (P = 0.004) but not histological grade or age. In addition, we found that 27% of the tumors showed LOH at 11q23.3-24 region between loci D11S934 and D11S912. Seven of nine MSI cases demonstrated low or no expression of p53. However, there was significantly reduced expression of PR, but not ER in MSI cases. CONCLUSIONS: Our results suggest that breast cancer acquires the RER phenotype (replication-error phenotype) in the relatively late stages, and that the RER phenotype is associated with aggressiveness of IDC (infiltrative duct carcinoma). The result also implicated that mismatch repair failure can alter the expression of PR but not ER and p53.