Differential sensitization of two human colon cancer cell lines to the antitumor effects of irinotecan combined with 5-aza-2'-deoxycytidine.

Irinotecan (CPT-11) is a key therapeutic drug used in the treatment of colorectal cancer, although acquired or constitutive resistance to CPT-11 (and its activated metabolite SN-38) can lead to tumor progression. Since the acquisition of drug resistance can result from DNA hypermethylation, the antitumor activity of CPT-11 and SN-38 was assessed in combination with a known DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine, also known as decitabine (DAC). DAC potentiated the antitumor activity of CPT-11 additively, and that of SN-38 synergistically, as measured by colony formation in the human colorectal cancer HCT116 cell line. No DAC potentiation of these antitumor effects was observed with another human colorectal cancer HT29 cell line. Anti-apoptotic B-cell lymphoma-2 (Bcl-2) protein expression was reduced to 50-67% of the control following a single treatment with CPT-11, SN-38, or DAC, and was markedly reduced to 7-8% following the combination of CPT-11/SN-38 with DAC. By contrast, Bcl-2 protein expression was barely detected in HT29. Wilms' tumor protein (WT1), which has been shown to be a positive regulator of Bcl-2 in HCT116 cells through WT1-kncokdown experiments, was downregulated in HCT116 and HT29 cells when treated with CPT-11/SN-38 combined with DAC, with decreases greater than any single administration of CPT-11, SN-38, or DAC. The extent of CPT-11/SN-38 potentiation by DAC may depend on Bcl-2 expression levels in human colorectal cancer cells.

Real-time ultrasound-guided infraorbital nerve block to treat trigeminal neuralgia using a high concentration of tetracaine dissolved in bupivacaine.

Background Trigeminal neuralgia is a neuropathic disorder characterized by episodes of intense pain in the face. Drug therapy is the first choice of treatment. However, in cases where drug therapy are contraindicated due to side effects, patients can get pain relief from lengthy neurosurgical procedures. Alternatively, a peripheral trigeminal nerve block can be easily performed in an outpatient setting. Therefore it is a useful treatment option for the acute paroxysmal period of TN in patients who cannot use drug therapy. We performed real-time ultrasound guidance for infraorbital nerve blocks in TN patients using a high concentration of tetracaine dissolved in bupivacaine. In this report, we examine the efficacy of our methods. Patients As approved by the Institutional Review Board, the medical records in our hospital were queried retrospectively. Six patients with TN at the V2 area matched the study criteria. All patients could not continue drug therapy with carbamazepine due to side effects and they received an ultrasound-guided infraorbital nerve block with a high concentration of tetracaine dissolved in bupivacaine. Methods The patient was placed in the supine position and the patient's face was sterilized and draped. An ultrasound system with a 6-13 MHz linear probe was used with a sterile cover. The probe was inserted into the horizontal plane of the cheek just beside the nose and was slid in the cranial direction to find the dimple of the infraorbital foramen. The 25G 25 mm needle was inserted from the caudal side just across from the probe using an out-of-plane approach. To lead the needle tip to the foramen, needle direction was corrected with real-time ultrasound guidance. After the test block with lidocaine (2%, 0.5 ml), a solution of tetracaine (20 mg) dissolved in bupivacaine (0.5%, 0.5 ml) was injected. During each injection, the spread of the agent around the nerve was confirmed using ultrasound images. Results Ten blocks were performed for six patients. Immediately after the procedure, all 10 blocks produced analgesia and relieved the pain. In the three blocks, pain was experienced in a new trigger point outside of the infraorbital nerve region (around the back teeth) within a week after the block and pain were relieved using other treatment. Two patients developed small hematomas in the cheek but they disappeared in a week. All patients did not complain about other side effects including paraesthesia, hyperpathia, dysaesthesia, or double vision. Hypoaesthesia to touch and pain in the infraorbital region were observed in all blocks after 2 weeks. Conclusions We performed real-time ultrasound-guided infraorbital nerve block for TN with a high concentration of tetracaine dissolved in bupivacaine. Our method achieved a high success rate and there were only minor and transient side effects. Implications Real-time ultrasound-guided infraorbital nerve block is one of the useful options to treat the acute paroxysmal period of TN at the infraorbital nerve area. Ultrasound-guided injections may become the standard practice for injecting peripheral trigeminal nerves. Using this high concentration of tetracaine as a neurolytic agent is effective and appears to have only minor side effects.

Sema4D stimulates axonal outgrowth of embryonic DRG sensory neurones.

Several semaphorins are thought to function as potent inhibitors of axonal growth. We have found that Sema4D stimulates axonal outgrowth of embryonic dorsal root ganglion (DRG) neurones in stead of retraction. Neutralizing antibodies to Sema4D inhibit this action. This action appears to differ slightly from that on PC12 cells, because DRG neurones respond to Sema4D without addition of nerve growth factor (NGF), while PC12 cells do not. On the other hand, it is blocked by deprivation of endogenous NGF with antibodies to NGF and also by Trk-inhibitor K252a, suggesting that endogenously produced-NGF and the activation of Trk receptor are required for Sema4D-action on DRG neurones. These indicate that neurite-outgrowth promoting actions of Sema4D are similar between DRG neurones and PC12 cells, since NGF-Trk signalling are required for these actions. Since Schwann cells can produce NGF, the contamination of these cells in our DRG culture might explain this action. In addition to plexin-B1 that is known as a Sema4D receptor, binding experiments indicate plexin-B2 as another receptor candidate for Sema4D. These plexins and Sema4D are expressed in embryonic DRGs. We suggest a new function of Sema4D as a neurite-outgrowth stimulating, autocrine/paracrine factor in embryonic sensory neurones.

Neurotrophic effect of Semaphorin 4D in PC12 cells.

Semaphorins provide crucial attractive and repulsive cues involved in axon guidance during neural development. Out of them, Semaphorin 4D (Sema4D) is enriched in the nervous and immune tissues, and acts as proliferative and survival factors of peripheral lymphocytes in the immune system, but is poorly understood in the nervous system. By using PC12 cells which are well known to differentiate into neural cells in response to nerve growth factor (NGF), we found that soluble forms of Sema4D had neurotrophic effects which were inhibited by neutralizing antibodies to Sema4D. Sema4D strikingly potentiated neurite outgrowth in the presence of 50 ng/ml NGF and increased sensitivity to NGF. Cells responded to very low concentrations of NGF in the presence of 1 nM Sema4D. Activation of following signal proteins, protein kinase C (PKC), L-type of voltage-dependent Ca(2+) channel, and phosphatidylinositol (PI) 3-kinase mediated neurotrophic neurite-outgrowth action of Sema4D. These findings suggest a new function of Sema4D as a neurotrophic signal in PC12 cells.