Polyethyleneimine- -Tocopherol Hydrogen Succinate/Hyaluronic Acid-Quercetin (PEI-TOS/HA-QU) Core-Shell Micelles Delivering Paclitaxel for Combinatorial Treatment of MDR Breast Cancer.

Multidrug resistance (MDR) remains a significant impediment to chemotherapy during cancer therapy. In this study, the amphiphilic biomaterials PEI-TOS and HA-QU were synthesized to self-assemble into PEI-TOS/HA-QU core-shell micelles for the targeted codelivery of paclitaxel (PTX) and quercetin (QU) to alleviate multidrug drug resistance and enhance therapeutic efficacy. The PTX-loaded micelles possessed a uniform particle size (167.60 ± 8.185 nm), stable negative charge (-19.13 ± 0.321 mV), and pH-responsive drug release with good compatibility. The drug-loaded micelles increased the chemosensitivity of MDR tumor cells (MDA-MB-231/MDR1) to PTX and activated mitochondria-dependent apoptotic pathways (the IC50 was 2.22-fold lower than that of PTX alone). Moreover, PEI-TOS/HA-QU micelles increased the cellular uptake of lipophilic antitumor drugs by downregulating P-gp expression in MDA-MB-231/MDR1 cells. Compared with Taxol, PTX-loaded PEI-TOS/HA-QU micelles presented excellent antitumor efficacy in tumor-bearing mice, with an average tumor size that was 3.7-fold lower than that of the control group. The drug-loaded formulation showed low in vitro / in vivo toxicity and better tumor accumulation than the free drug, which led to a high tumor inhibition rate of 80.56% and considerable biocompatibility. This work describes a new platform for the codelivery of lipophilic anticancer drugs and natural active ingredients such as PTX and QU for the treatment of MDR cancer cells.

Aristolochic acid I promoted clonal expansion but did not induce hepatocellular carcinoma in adult rats.

Aristolochic acid I (AAI) is a well-known nephrotoxic carcinogen, which is currently reported to be also associated with hepatocellular carcinoma (HCC). Whether AAI is a direct hepatocarcinogen remains controversial. In this study we investigated the association between AAI exposure and HCC in adult rats using a sensitive rat liver bioassay with several cofactors. Formation of glutathione S-transferase placental form-positive (GST-P+) foci was used as the marker for preneoplastic lesions/clonal expansion. We first conducted a medium-term (8 weeks) study to investigate whether AAI had any tumor-initiating or -promoting activity. Then a long-term (52 weeks) study was conducted to determine whether AAI can directly induce HCC. We showed that oral administration of single dose of AAI (20, 50, or 100 mg/kg) in combination with partial hepatectomy (PH) to stimulate liver proliferation did not induce typical GST-P+ foci in liver. In the 8-week study, only high dose of AAI (10 mg · kg-1 · d-1, 5 days a week for 6 weeks) in combination with PH significantly increased the number and area of GST-P+ foci initiated by diethylnitrosamine (DEN) in liver. Similarly, only high dose of AAI (10 mg· kg-1· d-1, 5 days a week for 52 weeks) in combination with PH significantly increased the number and area of hepatic GST-P+ foci in the 52-week study. No any nodules or HCC were observed in liver of any AAI-treated groups. In contrast, long-term administration of AAI (0.1, 1, 10 mg· kg-1· d-1) time- and dose-dependently caused death due to the occurrence of cancers in the forestomach, intestine, and/or kidney. Besides, AAI-DNA adducts accumulated in the forestomach, kidney, and liver in a time- and dose-dependent manner. Taken together, AAI promotes clonal expansion only in the high-dose group but did not induce any nodules or HCC in liver of adult rats till their deaths caused by cancers developed in the forestomach, intestine, and/or kidney. Findings from our animal studies will pave the way for further large-scale epidemiological investigation of the associations between AA and HCC.

[Differences of brain pathological changes and cognitive function after bilateral common carotid artery occlusion between Sprague-Dawley and Wistar rats].

The aim of the present study was to investigate the differences of the pathological changes and cognitive function after bilateral common carotid artery occlusion (BCCAO) between Sprague-Dawley (SD) and Wistar rats. Male SD and Wistar rats were randomly divided into 2 groups, respectively: sham operated (S-sham and W-sham) and operated (S-BCCAO and W-BCCAO) groups. The survival rate and the rate of loss of pupillary light reflex (PLR) were observed on day 1, 3, 7, 14 and 28 after the operation, and the light-dark box, Y-maze and odor recognition tests were performed to detect cognitive function on day 28 after the operation. HE and Luxol fast blue staining were used to observe the pathological changes of gray matter (hippocampus), white matter (optical tract), optic nerve, and retina. The results showed that the survival rate of the W-BCCAO group was 62.5%, and PLR loss rate was 100%; whereas the survival rate of the S-BCCAO group was 100%, and PLR loss rate was 58.3%. In the W-BCCAO group, percentages of time spent and distance traveled in the light box were more than those in the W-sham group, but there was no statistical significance between the S-BCCAO and S-sham groups. In the S-BCCAO group, the percentages of time spent and distance traveled in the III arm (labyrinth arm) of the Y-maze were less than those in the S-sham group, but no statistical significance was found between the W-BCCAO group and W-sham group. In the S-BCCAO group, the discrimination ratio of the odor recognition task was less than that in the S-sham group, but no statistical significance could be seen between the W-BCCAO and W-sham groups. Ischemic injury was observed in the CA1 area of the hippocampus in the S-BCCAO group, but no readily visible damage was observed in the W-BCCAO group. Ischemic injury of the visual beam and optic nerve was observed in both the S-BCCAO and W-BCCAO groups. Compared with the corresponding sham groups, the S-BCCAO and W-BCCAO groups showed serious retinal damage with significant thinner retina. The ganglion cell layer (GCL), inner plexiform layer (IPL), and outer plexiform layer (OPL) were thinner in the S-BCCAO group, but no statistical significances were shown in the other layers. All the layers, except the outer nuclear layer (ONL), were significantly thinner in the W-BCCAO group. The results indicate that there are differences of the pathological changes in the hippocampus and visual conduction pathway after BCCAO between SD and Wistar rats, and the degree of learning and memory injury was also different, which suggests that the vascular dementia model of different rat strains should be selected according to research purpose.

Plasma-specific microRNA response induced by acute exposure to aristolochic acid I in rats.

Aristolochic acid I (AAI) derived from a natural herbal alkaloid is a nephrotoxicant. AAI-induced acute kidney injury (AKI), a devastating clinical disease associated with high mortality rates, is difficult for early diagnosis. To address this issue, we identified and validated early-detection biomarkers for AAI-induced acute kidney injury via profiling microRNA expression in rats. Global miRNA expression profile analysis found that 21 miRNAs were significantly dysregulated in kidney of rats treated by 40 mg/kg AAI on day 2, day 4, or day 6, among which 5 miRNAs were upregulated at all three time points. Quantitative RT-PCR confirmed that miR-21-3p on day 4 and day 6 was obviously upregulated in kidney of rats treated by 40 mg/kg AAI. Further examination found that miR-21-3p was increased in plasma early on day 2 in 10 mg/kg AAI-treated rats, but not in non-target organs. Importantly, the elevation of plasma miR-21-3p preceded the increase in blood urea nitrogen and creatinine, and the presence of renal tubular injury, characterized by differential increase before and after the presence of renal tubular lesions. Our findings thus show that miRNA expression is upregulated in kidney and plasma of AKI rat induced by AAI, and plasma miR-21-3p may be served as a new potential biomarker for early diagnosing AAI-induced acute kidney injury in rats, and possibly in humans.

Oral exposure to aristolochic acid I induces gastric histological lesions with non-specific renal injury in rat.

Many Aristolochia species herbal drugs, used for diseases treatment since antiquity, contain active component aristolochic acid mixture, which consists of aristolochic acid I and II. However, it remains unclear whether aristolochic acid I is gastrotoxic, though evidence has shown that aristolochic acid mixture is nephrotoxic, carcinogenic, and genotoxic. The present study aimed to investigate the gastrotoxicity in rats treated with aristolochic acid I alone. Four groups of rats were orally administrated with vehicle (1% NaHCO3), or 30mg, 60mg, and 90mg/kg/day of aristolochic acid I for twelve days. The results showed that aristolochic acid I can induce obvious body weight loss, forestomach injury characterized by necrosis, ulcer, hyperkeratosis, and hyperplasia of epithelial cells. The severity of these forestomach lesions was presented in a dose-dependent mode. Meanwhile, only non-specific, slight renal tubule degeneration, and occasionally single necrotic epithelial cell were found in aristolochic acid I-treated rats' kidney. These resulst indicated aristolochic acid I had obvious gastrotoxicity, and such aristolochic acid I-induced forestomach toxicity probably presented much prior to kidney injury. Such irritation lesions may play a partial role in gastric cancer development of rats induced by aristolochic acid. Therefore, these results expanded our understanding on the digestive system toxicity of aristolochic acid I.

Dynamics of global gene expression changes during brain metastasis formation.

As methods of cancer diagnosis and treatment improve, interest in metastatic brain tumors continues to increase. In the present study, we attempted to characterize genetically the dynamic changes occurring during brain metastasis formation by DNA microarray, and attempted to compare these findings with histological observations. Lewis lung carcinoma cells were injected into C57BL/6Ncrj mice carotid arteries. The mice were sacrificed at days 1-9 after injection. We performed histological observation and genome-wide expression profiling using a DNA microarray. In histological observation, tumor cells were observed in capillary vessels at day 1 after injection. At day 3, the tumor cells had begun to proliferate. At day 6, the metastatic foci showed "perivascular proliferations". Next, we performed a pairwise comparison of gene expression microarray data from day 1 to day 9 after injection. The first major change occurred between Phase Two and Phase Three. When hierarchical clustering was performed between different samples using the 867 genes, they could be classified into identical clusters for days 1 and 2, identical clusters for day 3 to day 5, and identical clusters for day 6 to day 9. For time course analysis, we extracted 623 genes by the pairwise comparison. By using the quality threshold (QT) nonhierarchical clustering method, we identified 37 expression patterns. These patterns can be separated into eight clusters by using the k-means method. The microarray results reported here strongly suggest that a large number of genes exhibit a spike pattern, which is tantamount to phase-specific expression.

Comparison of metastatic brain tumour models using three different methods: the morphological role of the pia mater.

As methods of cancer diagnosis and treatment progress, interest in metastatic brain tumours continues to increase. There are many studies using various methods of animal model and we considered that each model reflects different pathological processes because of the unique composition of the brain. We prepared metastatic brain tumour models using three different methods. In this study, we attempted to elucidate the roles of the pia mater in brain metastasis. The metastatic foci showed an angiocentric pattern, forming collars of neoplastic cells, and were designated 'perivascular proliferations'. Furthermore, we observed neoplastic cells that infiltrated the brain parenchyma, the border of which had become indistinct. These were labelled 'invasive proliferations'. The internal carotid artery injection model reflects haematogenous metastasis. In this model, both perivascular and invasive proliferations were observed. The intrathecal injection model reflects leptomeningeal carcinomatosis. In this model, metastasis to the meninges was observed. In the stereotactic injection model, the tumour proliferation at the injection site and the infiltration into the brain parenchyma were observed. The pia-glial membrane serves as a scaffold when neoplastic cells spread to the perivascular space forming angiocentric pattern. The pia-glial membrane is found between the brain parenchyma and blood vessels. Blood vessels penetrate the brain through tunnels known as perivascular spaces that are covered by pia mater. Three different methods which we prepared reflect three different pathological processes. Our findings suggest that the pia mater is a critical factor in brain metastasis.