The androgens and androgen receptor (AR) play key roles in the prostate cancer (PCa) development and progression via epithelium-stroma cross talk. Prostate cancer-associated fibroblasts (CAFs) are dominant components in PCa stroma and are essential in the malignant progression by supporting tumorigenesis and metastasis. However, the AR roles in CAFs are still obscure. We isolated and immortalized the CAFs from human PCa tissues and found the CAFs are AR positive. We then knocked down their AR with siRNA and co-cultured the resultant CAFs with PCa cell line PC3. The MTT, invasion, and colony formation assays were performed to study the PC3 biological behavior. The results showed that the PCa epithelial growth, invasion, and colony formation abilities decreased when knocking down the CAFs AR. By using the real-time quantitative polymerase chain reaction, we found the IGF1, FGF7, FGF10, SDF1, HGF, and TGFb2 expression levels decreased in the AR knocked down CAFs. [1]
The CAFs and NPFs were successfully cultured, immortalized, and characterized. The CAFs were able to transform prostate epithelial cells into malignant cells, but NPFs were not. The CAFs were more active in promoting proliferation of and invasion by PCa cells, and in secreting higher levels of a series of growth factors. The immortalized CAFs were more supportive of PCa carcinogenesis and progression. Targeting CAFs might be a potential option for PCa therapy. Immortalized CAFs and NPFs will also be valuable resources for future experimental exploration. Focused ultrasound (FUS) has become an important non-invasive therapy for prostate tumor ablation via thermal effects in the clinic. The cavitation effect induced by FUS is applied for histotripsy, support drug delivery, and the induction of blood vessel destruction for cancer therapy. Numerous studies report that cavitation-induced sonoporation could provoke multiple anti-proliferative effects on cancer cells. Therefore, cavitation alone or in combination with thermal treatment is of great interest but research in this field is inadequate. [2]
Human prostate cancer cells (LNCap and PC-3) were exposed to 40 s cavitation using a FUS system, followed by water bath hyperthermia (HT). The clonogenic assay, WST-1 assay, and Transwell® invasion assay, respectively, were used to assess cancer cell clonogenic survival, metabolic activity, and invasion potential. Fluorescence microscopy using propidium iodide (PI) as a probe of cell membrane integrity was used to identify sonoporation. The H2A.X assay and Nicoletti test were conducted in the mechanism investigation to detect DNA double-strand breaks (DSBs) and cell cycle arrest. Immunofluorescence microscopy and flow cytometry were performed to determine the distribution and expression of 5α-reductase.
Short FUS shots with cavitation (FUS-Cav) in combination with HT resulted in, respectively, a 2.2, 2.3, and 2.8-fold decrease (LNCap) and a 2.0, 1.5, and 1.6-fold decrease (PC-3) in the clonogenic survival, cell invasiveness and metabolic activity of prostate cancer cells when compared to HT alone. FUS-Cav immediately induced sonoporation in 61.7% of LNCap cells, and the combination treatment led to a 1.4 (LNCap) and 1.6-fold (PC-3) increase in the number of DSBs compared to HT alone. Meanwhile, the combination therapy resulted in 26.68% of LNCap and 31.70% of PC-3 with cell cycle arrest in the Sub-G1 phase and 35.37% of PC-3 with cell cycle arrest in the G2/M phase. Additionally, the treatment of FUS-Cav combined with HT block the androgen receptor (AR) signal pathway by reducing the relative Type I 5α-reductase (SRD5A1) level to 38.28 ± 3.76% in LNCap cells, and decreasing the relative Type III 5α-reductase 3 (SRD5A3) level to 22.87 ± 4.88% in PC-3 cells, in contrast, the relative SRD5A level in untreated groups was set to 100%.
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