Mat-LyLu Rat Prostate Cancer Cell Research Articles

Cancer pain and neuropathic pain are associated with A β sensory neuronal plasticity in dorsal root ganglia and abnormal sprouting in lumbar spinal cord.

Evidence suggests that there are both nociceptive and neuropathic components of cancer-induced pain. We have observed that changes in intrinsic membrane properties and excitability of normally non-nociceptive Aβ sensory neurons are consistent in rat models of peripheral neuropathic pain and cancer-induced pain. This has prompted a comparative investigation of the intracellular electrophysiological characteristics of sensory neurons and of the ultrastructural morphology of the dorsal horn in rat models of neuropathic pain and cancer-induced pain. Neuropathic pain model rats were induced with a polyethylene cuff implanted around a sciatic nerve. Cancer-induced pain model rats were induced with mammary rat metastasis tumour-1 rat breast cancer or MATLyLu rat prostate cancer cells implanted into the distal epiphysis of a femur. Behavioural evidence of nociception was detected using von Frey tactile assessment. Aβ-fibre low threshold mechanoreceptor neurons in both cancer-induced pain and neuropathic pain models exhibited slower dynamics of action potential genesis, including a wider action potential duration and lower action potential amplitude compared to those in control animals. Enhanced excitability of Aβ-fibre low threshold mechanoreceptor neurons was also observed in cancer-induced pain and neuropathic pain models. Furthermore, both cancer-induced pain and neuropathic pain models showed abundant abnormal axonal sprouting in bundles of myelinated axons in the ipsilateral spinal laminae IV and V. The patterns of changes show consistency between rat models of cancer-induced pain and neuropathic pain. These findings add to the body of evidence that animal models of cancer-induced pain and neuropathic pain share features that may contribute to the peripheral and central sensitization and tactile hypersensitivity in both pain states.

9P - Anti-metastatic effect of riluzole on Mat-LyLu rat prostate cancer cell line

9P - Anti-metastatic effect of riluzole on Mat-LyLu rat prostate cancer cell line

Rat model of cancer-induced bone pain: changes in nonnociceptive sensory neurons in vivo.

Clinical data on cancer-induced bone pain (CIBP) suggest extensive changes in sensory function. In a previous investigation of an animal model of CIBP, we have observed that changes in intrinsic membrane properties and excitability of dorsal root ganglion (DRG) nociceptive neurons correspond to mechanical allodynia and hyperalgesia. To investigate the mechanisms underlying changes in nonnociceptive sensory neurons in this model, we have compared the electrophysiological properties of primary nonnociceptive sensory neurons at <1 and >2 weeks after CIBP model induction with properties in sham control animals. Copenhagen rats were injected with 106 MAT-LyLu rat prostate cancer cells into the distal femur epiphysis to generate a model of CIBP. After von Frey tactile measurement of mechanical withdrawal thresholds, the animals were prepared for acute electrophysiological recordings of mechanically sensitive neurons in the DRG in vivo. The mechanical withdrawal threshold progressively decreased in CIBP model rats. At <1 week after model induction, there were no changes observed in nonnociceptive Aβ-fiber DRG neurons between CIBP model rats and sham rats. However, at >2 weeks, the Aβ-fiber low-threshold mechanoreceptors (LTMs) in CIBP model rats exhibited a slowing of the dynamics of action potential (AP) genesis, including wider AP duration and lower AP amplitude compared with sham rats. Furthermore, enhanced excitability of Aβ-fiber LTM neurons was observed as an excitatory discharge in response to intracellular injection of depolarizing current into the soma. After induction of the CIBP model, Aβ-fiber LTMs at >2 weeks but not <1 week had undergone changes in electrophysiological properties. Importantly, changes observed are consistent with observations in models of peripheral neuropathy. Thus, Aβ-fiber nonnociceptive primary sensory neurons might be involved in the peripheral sensitization and tumor-induced tactile hypersensitivity in CIBP.

Activity‐dependent regulation of voltage‐gated Na+ channel expression in Mat‐LyLu rat prostate cancer cell line

We have shown previously that voltage-gated Na(+) channels (VGSCs) are up-regulated in human metastatic disease (prostate, breast and small-cell lung cancers), and that VGSC activity potentiates metastatic cell behaviours. However, the mechanism(s) regulating functional VGSC expression in cancer cells remains unknown. We investigated the possibility of activity-dependent (auto)regulation of VGSC functional expression in the strongly metastatic Mat-LyLu model of rat prostate cancer. Pretreatment with tetrodotoxin (TTX) for 24-72 h subsequently suppressed peak VGSC current density without affecting voltage dependence. The hypothesis was tested that the VGSC auto-regulation occurred via VGSC-mediated Na(+) influx and subsequent activation of protein kinase A (PKA). Indeed, TTX pretreatment reduced the level of phosphorylated PKA, and the PKA inhibitor KT5720 decreased, whilst the adenylate cyclase activator forskolin and the Na(+) ionophore monensin both increased the peak VGSC current density. TTX reduced the mRNA level of Nav1.7, predominant in these cells, and VGSC protein expression at the plasma membrane, although the total VGSC protein level remained unchanged. TTX pretreatment eliminated the VGSC-dependent component of the cells' migration in Transwell assays. We concluded that the VGSC activity in Mat-LyLu rat prostate cancer cells was up-regulated in steady-state via a positive feedback mechanism involving PKA, and this enhanced the cells' migratory potential.

Isoliquiritigenin inhibits migration and invasion of prostate cancer cells

Isoliquiritigenin (ISL), 4,2′,4′-trihydroxychalcone, found in licorice, shallot, and bean sprouts, is a potent antioxidant with anti-inflammatory, anti-platelet aggregation and anti-carcinogenic effects. The purpose of the present study was to investigate the effects of ISL treatment on the migration and invasion characteristics of DU145 prostate cancer cells. DU145 cells were cultured in the presence of 0 – 20 μmol/L ISL with 10μng/mL EGF, 10 nmol/L IGF-I, or 20 ng/mL heregulin-β. EGF stimulated migration, whereas IGF-I or hergulin had no effect. ISL inhibited the basal and EGF-induced cell migration and invasion in dose-dependent manners. Gelatin zymography and Western blot analysis exhibited a significant down-regulation of matrix metalloproteinase (MMP)-9 expression in DU145 cells stimulated with phorbol 12-myristate 13-aceate (PMA). ISL decreased the protein levels of tissue inhibitor of metalloproteinase-1 (TIMP-1) but increased TIMP-2 levels in DU145 cells in a concentration-dependent manner. In MatLyLu rat prostate cancer cells, ISL suppressed the protein levels of both pro and active MMP-2 and -9 and mRNA levels of MMP-2. These data therefore provide direct evidence for the role of ISL as a potent antimetastatic agent, which can markedly inhibit the metastatic and invasive capacity of malignant cells. The decrease in MMP-9 and increase in TIMP-2 expression may be one of mechanisms by which ISL inhibits cancer cell invasion.

Serum concentration modifies amplitude and kinetics of voltage-gated Na + current in the Mat-LyLu cell line of rat prostate cancer

Voltage-gated Na + channel (VGSC) expression has previously been shown to be upregulated in strongly metastatic prostate cancer cells (rat and human) and its activity shown to potentiate a variety of cellular behaviours integral to the metastatic cascade. However, the mechanism(s) responsible for the Na + channel upregulation is not known. As a step towards evaluating the role of the extracellular biochemical environment in this regard, we have determined the effects of serum concentration on characteristics of Na + channel expressed in the strongly metastatic Mat-LyLu rat prostate cancer cell line. Whole-cell patch-clamp recording techniques were used to study the effects of serum concentrations, above and below the normal 1%. Both the amplitude and the kinetics of the currents were analysed. The following results were obtained: (1) Adding 1% foetal calf serum to cells starved of serum for 24 h increased Na + current density; however, increasing serum concentration further (to 5%) caused a reduction. (2) Serum-free medium produced Na + currents with slower kinetics of activation (time to peak) and inactivation (exponential decay). (3) Increased serum concentration (a) shifted steady-state inactivation to more positive potentials without affecting conductance and (b) increased tetrodotoxin sensitivity. It is concluded that serum concentration is an important determinant of the Na + channel characteristics leading to possible transcriptional and post-translational modifications of channel expression and/or activity. Experiments are now needed to determine which constituents (protein hormones, growth factors, etc.) are responsible for these effects.

The Growth Regulatory Fibroblast IK Channel Is the Prominent Electrophysiological Feature of Rat Prostatic Cancer Cells

Physiological effectors for mitogenic cell growth control remain to be determined for mammalian tumor cells, particularly those derived from prostatic tissue. One such effector for mitogenic Ras/MAPK signaling in fibroblasts is an intermediate-conductance, calcium-activated potassium channel (FIK). In this study patch-clamp electrophysiology was used to show that both AT2.1 and MatLyLu rat prostate cancer cell lines express high levels of a current identified as FIK, based on the following criteria: activation by elevation of intracellular calcium, voltage independence, potassium selectivity, and block by charybdotoxin (ChTX) and the Stichodactyla helianthus potassium channel neurotoxin (StK). FIK current densities in AT2.1 and MatLyLu cells were comparable to the high levels seen in fibroblasts transfected with oncogenic Ras or Raf, suggesting hyperactivity of the Ras/MAPK pathway in prostatic cancer cells. Voltage-gated sodium current was present in most MatLyLu cells but absent from AT2.1 cells, and all AT2.1 cells had voltage-gated potassium currents. Thus, FIK is the main electrophysiological feature of rat prostatic cancer cells as it is for mitogenically active fibroblasts, suggesting it may play a similar growth regulatory role in both.

Tetrodotoxin suppresses morphological enhancement of the metastatic MAT-LyLu rat prostate cancer cell line.

Voltage-gated Na+ channels are expressed by highly metastatic MAT-LyLu cells, but not by poorly metastatic AT-2 cells, derived from the rodent Dunning model of prostatic cancer. We have investigated the possible involvement of these channels in the morphological development of the cells. Incubation of both the MAT-LyLu and the AT-2 cell line for 24 h with the Na+ channel blocker tetrodotoxin (TTX) at 6 microM altered the morphology only of the MAT-LyLu cell line. TTX produced significant decreases in: (a) cell process length and (b) field diameter, and increases in (c) cell body diameter and (d) process thickness. Importantly, 6 microM TTX had no significant effects on proliferation rates or cellular toxicity. The results suggest that Na+ channel activity plays a significant role in determining the morphological development of MAT-LyLu cells in such a way as to enhance their metastatic potential.

Electrophysiological characterization of voltage-gated Na+ current expressed in the highly metastatic Mat-LyLu cell line of rat prostate cancer.

Voltage-gated Na+ channels, classically associated with impulse conduction in excitable tissues, are also found in a variety of epithelial cell types where their possible functions are not known so well. We have previously reported expression of a voltage-gated Na+ channel specifically in the highly metastatic Mat-LyLu rat prostate cancer cell line; blockage of the current with tetrodotoxin (TTX) significantly reduced the invasiveness of the cells in vitro, suggesting that the channel may have a functional role in metastasis. The aim of the present study was to characterize this current using the whole-cell patch clamp recording technique, and compare it to Na+ currents found in various other tissues. The inward current of the Mat-LyLu cells was abolished completely, but reversibly, in Na+-free solution, confirming that Na+ was indeed the permeant ion. Activation occurred at -40 mV and currents reached a maximal amplitude at around 6 mV. Boltzmann fits to current activation and steady-state inactivation revealed that the currents were half activated at about -15 mV and half inactivated at -80 mV. Both current inactivation and recovery from inactivation followed a double-exponential time course with fast and slow components. The Na+ currents were highly sensitive to block by TTX (IC50 approximately 18 nM), whilst 1 microM mu-conotoxin GIIIA mostly had no effect. 100 microM Cd2+ also had no effect on the current, whilst 2.5 mM Cd2+, Mn2+, and Co2+ each caused a depolarizing shift in activation and a reduction in peak conductance of around 20%. In conclusion, the Na+ channel expressed in the highly metastatic Mat-LyLu cell line appeared to have electrophysiological and pharmacological properties of TTX-sensitive channels. Further work is needed, however, to elucidate the exact nature of the channel protein and the mechanism(s) of its involvement in cellular invasiveness.

Effects of transforming growth factor beta 1 on the adenylyl cyclase-cAMP pathway in prostate cancer.

We reported previously that MATLyLu rat prostate cancer cells engineered to overproduce transforming growth factor beta 1 (TGF beta 1) produce larger, more metastatic tumors in vivo. We recognized that this ability of TGF beta 1 to act as a positive modulator of prostate tumor behavior might be due to effects of TGF beta 1 on the host and/or on the tumor cells. In this study we demonstrated that the cells themselves respond to endogenously produced TGF beta 1, and that the adenylyl cyclase (AC)-cAMP pathway is affected. TGF beta 1-overproducing cells had lower membrane AC activity, lower intracellular cAMP content, and a lower Gs alpha protein level than did control cells. Prostate cancer cells were growth inhibited by 8-bromo-cAMP or forskolin, agents that elevate intracellular cAMP. Thus, TGF beta 1 overproduction affects the phenotype of the tumor cells, deliberate activation of endogenously produced latent TGF beta 1 is not required (indicating that the cells themselves are capable of activating latent TGF beta 1), and TGF beta 1 overproduction lowers the cellular concentration of the growth inhibitor cAMP. Therefore, TGF beta 1 overproduction could affect tumor behavior in vivo in part via a direct effect on the tumor cells.