Posttraumatic stress disorder: from diagnosis to prevention.

Posttraumatic stress disorder (PTSD) is a chronic impairment disorder that occurs after exposure to traumatic events. This disorder can result in a disturbance to individual and family functioning, causing significant medical, financial, and social problems. This study is a selective review of literature aiming to provide a general outlook of the current understanding of PTSD. There are several diagnostic guidelines for PTSD, with the most recent editions of the DSM-5 and ICD-11 being best accepted. Generally, PTSD is diagnosed according to several clusters of symptoms occurring after exposure to extreme stressors. Its pathogenesis is multifactorial, including the activation of the hypothalamic-pituitary-adrenal (HPA) axis, immune response, or even genetic discrepancy. The morphological alternation of subcortical brain structures may also correlate with PTSD symptoms. Prevention and treatment methods for PTSD vary from psychological interventions to pharmacological medications. Overall, the findings of pertinent studies are difficult to generalize because of heterogeneous patient groups, different traumatic events, diagnostic criteria, and study designs. Future investigations are needed to determine which guideline or inspection method is the best for early diagnosis and which strategies might prevent the development of PTSD.

DNMT3a contributes to the development and maintenance of bone cancer pain by silencing Kv1.2 expression in spinal cord dorsal horn.

Abstract: Metastatic bone tumor-induced changes in gene transcription and translation in pain-related regions of the nervous system may participate in the development and maintenance of bone cancer pain. Epigenetic modifications including DNA methylation regulate gene transcription. Here, we report that intrathecal injection of decitabine, a DNA methyltransferase (DNMT) inhibitor, dose dependently attenuated the development and maintenance of bone cancer pain induced by injecting prostate cancer cells into the tibia. The level of the de novo DNMT3a, but not DNMT3b, time dependently increased in the ipsilateral L4/5 dorsal horn (not L4/5 dorsal root ganglion) after prostate cancer cells injection. Blocking this increase through microinjection of recombinant adeno-associated virus 5 (AAV5) expressing Dnmt3a shRNA into dorsal horn rescued prostate cancer cells-induced downregulation of dorsal horn Kv1.2 expression and impaired prostate cancer cells-induced pain hypersensitivity. In turn, mimicking this increase through microinjection of AAV5 expressing full-length Dnmt3a into dorsal horn reduced dorsal horn Kv1.2 expression and produced pain hypersensitivity in the absence of prostate cancer cells injection. Administration of neither decitabine nor virus affected locomotor function and acute responses to mechanical, thermal, or cold stimuli. Given that Dnmt3a mRNA is co-expressed with Kcna2 mRNA (encoding Kv1.2) in individual dorsal horn neurons, our findings suggest that increased dorsal horn DNMT3a contributes to bone cancer pain through silencing dorsal horn Kv1.2 expression. DNMT3a may represent a potential new target for cancer pain management.

Trypsin-protease activated receptor-2 signaling contributes to pancreatic cancer pain.

Pain treatment is a critical aspect of pancreatic cancer patient clinical care. This study investigated the role of trypsin-protease activated receptor-2 (PAR-2) in pancreatic cancer pain. Pancreatic tissue samples were collected from pancreatic cancer (n=22) and control patients (n=22). Immunofluorescence analyses confirmed colocalization of PAR-2 and neuronal markers in pancreatic cancer tissues. Trypsin levels and protease activities were higher in pancreatic cancer tissue specimens than in the controls. Supernatants from cultured human pancreatic cancer tissues (PC supernatants) induced substance P and calcitonin gene-related peptide release in dorsal root ganglia (DRG) neurons, and FS-NH2, a selective PAR-2 antagonist, inhibited this effect. A BALB/c nude mouse orthotopic tumor model was used to confirm the role of PAR-2 signaling in pancreatic cancer visceral pain, and male Sprague-Dawley rats were used to assess ambulatory pain. FS-NH2 treatment decreased hunch scores, mechanical hyperalgesia, and visceromotor reflex responses in tumor-bearing mice. In rats, subcutaneous injection of PC supernatant induced pain behavior, which was alleviated by treatment with FS-NH2 or FUT-175, a broad-spectrum serine protease inhibitor. Our findings suggest that trypsin-PAR-2 signaling contributes to pancreatic cancer pain in vivo. Treatment strategies targeting PAR-2 or its downstream signaling molecules might effectively relieve pancreatic cancer pain.

Suppression of acute morphine withdrawal syndrome by adenovirus-mediated ß-endorphin in rats.

BACKGROUND: Endogenous ß-endorphin (ß-EP) in the central nervous system (CNS) is decreased upon opioid addiction. The current study examined whether exogenous ß-EP, delivered using an adenoviral vector into the CNS could attenuate morphine withdrawal syndrome in rats. METHODS: The model of opioid-dependent rats was set up by receiving subcutaneous injection of morphine using an escalating regimen for 6days (5, 10, 20, 40, 50, 60mg/kg, three times/day). The adenovirus mediated ß-EP gene was constructed based on our previous work. The ilea of opioid-dependent rats were isolated and treated with the supernatant of Ad-NEP. The basic and naloxone-induced (4µm/l) contractions of dependent ilea were recorded. The Ad-NEP was injected into the left lateral ventricle of the addition rats. The expression of the ß-EP gene was verified by radioimmunoassay of the cerebrospinal fluid (CSF) and immunocytochemistry for ß-EP. Withdrawal syndrome was evaluated after intraperitoneal injection of naloxone. RESULTS: The contractions of dependent ilea were attenuated with supernatant containing ß-EP expressed by Ad-NEP. Injection of the Ad-NEP resulted in significant increases in ß-EP level in the CSF and ß-EP-positive neurons. Rats receiving adenovirus carrying the ß-EP gene had significantly less severe withdrawal symptoms upon naloxone challenge. CONCLUSIONS: Exogenous ß-EP mediated by adenovirus could attenuate withdrawal syndrome in morphine-dependent rats.

Intrathecal siRNA against Toll-like receptor 4 reduces nociception in a rat model of neuropathic pain.

BACKGROUND: Neuropathic pain is characterized by hyperalgesia, allodynia and spontaneous pain. It often occurs as a result of injury to peripheral nerves, dorsal root ganglions (DRG), spinal cord, or brain. Recent studies have suggested that Toll-like receptor 4 (TLR4) might play a role in neuropathic pain. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we investigated the role of TLR4 in a rat chronic constriction injury (CCI) model and explored the feasibility of treating neuropathic pain by inhibiting TLR4. Our results demonstrated that intrathecal siRNA-mediated suppression of TLR4 attenuated CCI-induced mechanical allodynia and thermal hyperalgesia through inhibiting the activation of NF-kappaB p65 and production of proinflammatory cytokines (e.g., TNF-alpha and IL-1 beta). CONCLUSIONS/SIGNIFICANCE: These findings suggest that suppression of TLR4 mediated by intrathecally administered siRNA may be a new strategy for the treatment of neuropathic pain.

Bilateral downregulation of Nav1.8 in dorsal root ganglia of rats with bone cancer pain induced by inoculation with Walker 256 breast tumor cells.

BACKGROUND: Rapid and effective treatment of cancer-induced bone pain remains a clinical challenge and patients with bone metastasis are more likely to experience severe pain. The voltage-gated sodium channel Nav1.8 plays a critical role in many aspects of nociceptor function. Therefore, we characterized a rat model of cancer pain and investigated the potential role of Nav1.8. METHODS: Adult female Wistar rats were used for the study. Cancer pain was induced by inoculation of Walker 256 breast carcinosarcoma cells into the tibia. After surgery, mechanical and thermal hyperalgesia and ambulation scores were evaluated to identify pain-related behavior. We used real-time RT-PCR to determine Nav1.8 mRNA expression in bilateral L4/L5 dorsal root ganglia (DRG) at 16-19 days after surgery. Western blotting and immunofluorescence were used to compare the expression and distribution of Nav1.8 in L4/L5 DRG between tumor-bearing and sham rats. Antisense oligodeoxynucleotides (ODNs) against Nav1.8 were administered intrathecally at 14-16 days after surgery to knock down Nav1.8 protein expression and changes in pain-related behavior were observed. RESULTS: Tumor-bearing rats exhibited mechanical hyperalgesia and ambulatory-evoked pain from day 7 after inoculation of Walker 256 cells. In the advanced stage of cancer pain (days 16-19 after surgery), normalized Nav1.8 mRNA levels assessed by real-time RT-PCR were significantly lower in ipsilateral L4/L5 DRG of tumor-bearing rats compared with the sham group. Western-blot showed that the total expression of Nav1.8 protein significantly decreased bilaterally in DRG of tumor-bearing rats. Furthermore, as revealed by immunofluorescence, only the expression of Nav1.8 protein in small neurons down regulated significantly in bilateral DRG of cancer pain rats. After administration of antisense ODNs against Nav1.8, Nav1.8 protein expression decreased significantly and tumor-bearing rats showed alleviated mechanical hyperalgesia and ambulatory-evoked pain. CONCLUSIONS: These findings suggest that Nav1.8 plays a role in the development and maintenance of bone cancer pain.

Acute PAR2 activation reduces alpha, beta-MeATP sensitive currents in rat dorsal root ganglion neurons.

It has been reported that proteinase-activated receptor 2 (PAR2) receptor activation enhances the animal's pain response and PAR2 coexpresses with P2X3 in dorsal root ganglion neurons. However, whether PAR2 activation has a direct impact on P2X3 currents is still not clear. In this study, we performed the patch-clamp experiments in cultured dorsal root ganglion neurons and found that when incubated with trypsin or the PAR2 agonist SL-NH2 for a short time (3 min), instead of increasing, P2X3 currents amplitude decreased significantly. Meanwhile, the opening of P2X3 ion channel accelerated. Protein kinase A inhibitor H89 could not reverse above phenomenon, but played a synergistic effect on the contrary. These results suggest that the enhanced pain response caused by PAR2 activation is not through direct increase of the P2X3 current amplitude, and the acceleration of P2X3 opening may participate in the enhanced pain response in a long-time view. Moreover, protein kinase A does not participate in the inhibition of P2X3 currents caused by PAR2 activation.

Human signal peptide had advantage over mouse in secretory expression.

The signal peptide is a critical component in the secretory expression of protein in eukaryotic cells. It has been verified that the signal peptide of mouse nerve growth factor could mediate the secretory expression of beta-endorphin in cultured non-neuronal cells. Although there is a counterpart of nerve growth factor in human genome, no research about the signal sequence from human genome has been reported. The function of mediating secretory expression is affected by many factors. We assumed that the counterpart from human genome could function as the signal peptide from mouse nerve growth factor does and these two signal sequences had different efficiency in mediating secretory expression of beta-endorphin, but we could not figure out which one had a better function. To validate our hypothesis and give an answer to the question, we constructed two eukaryotic vectors, pcDNA3.1-hEP and pcDNA3.1-mEP, containing human and mouse signal sequences in fusion genes, respectively. RT-PCR showed that the constructed fusion genes were expressed in NIH3T3 cells. We also found that the detected beta-endorphin by the immunofluorescent technique was mainly in the cytoplasm of NIH3T3 cells. The concentration of beta-endorphin in the culture medium by RIA is 280.33 +/- 24.16 (pg/ml) and 191.04 +/- 7.96 (pg/ml) from pcDNA3.1-hEP and pcDNA3.1-mEP, respectively, and there was a significant statistical difference between them (P < 0.05). A difference existed between them and that from blank vector individually (P < 0.01). These findings suggest that our constructed fusion gene containing the signal sequence of human nerve growth factor can be secretorily expressed and the efficiency of the signal peptide from human nerve growth factor is higher than that of mouse signal peptide.