Effects of intrathecal injection of resveratrol on sense of pain and OPG/RANK/RANK-L signal pathway in rats with bone cancer pain / 中国药房

OBJECTIVE To study the effects of intrathecal injection of resveratrol on sense of pain and OPG/RANK/RANK-L signal pathway in rats with bone cancer pain. METHODS After treatment via intrathecal tube ，the rats were randomly divided into sham operation group ，model group ，resveratrol low-dose ，medium-dose and high-dose groups （0.25，1，2 mg/kg），with 8 rats in each group. Except for sham operation group ，other groups were given Walker 256 breast cancer cell suspension on the right hind limb tibia to induce bone cancer pain model. After 12 days of modeling ，intrathecal administration was performed in resveratrol groups. The model group and sham operation group received the same volume of dimethyl sulfoxide ，10 μL，once a day ，for 5 days. The paw withdrawal mechanical threshold （PWMT）and paw withdrawal thermal latency （PWTL）were measured at 3 d，5 d after administration. The serum levels of tumor necrosis factor α（TNF-α），interleukin 6（IL-6），IL-1β and chemokine ligand 2 （CCL2）were determined after last medication ，and the protein expressions of RANK ，RANK-L and OPG in tibia were also determined. RESULTS Compared with sham operation group ，PWMT and PWTL were significantly decreased in the model group 12 days after operation （P＜0.05）. Compared with model group ，PWMT and PWTL were significantly increased in resveratrol medium-dose and high-dose groups at 3 d and 5 d after administration （P＜0.05）. Compared with model group ，the levels of TNF-α，IL-6，IL-1β and CCL2 in serum were significantly decreased in resveratrol middle-dose and high-dose groups at 5 d after administration（P＜0.05）；the expression of IL- 6 in serum was significantly decreased in resveratrol low-dose group （P＜0.05）； the protein expressions of RANK-L and RANK in tibia were significantly decreased ，while the protein expression of OPG was increased significantly in resveratrol middle-dose group （P＜0.05）. CONCLUSIONS Resveratrol by intrathecal injection can relieve the inflammation caused by bone cancer pain of rats ， the mechanism of which may be associated with inhibiting the activity of OPG/RANK/RANK-L signal pathway.

Early intervention of bone-nearby acupuncture combined with electroacupuncture on morphine tolerance in bone cancer pain rats and its effect on the expression of HDAC and MOR in dorsal root ganglia / 中国针灸

OBJECTIVE@#To observe the effect of early intervention of bone-nearby acupuncture (BNA) combined with electroacupuncture (EA) on the expression of histone deacetylase1(HDAC1), histone deacetylase 2 (HDAC2) andμ-opioid recepter (MOR) in dorsal root ganglia (DRG) of bone cancer pain-morphine tolerance (BCP-MT) rats, and to explore its possible mechanism.@*METHODS@#A total of 35 SD rats were randomized into a sham BCP group (=6), a BCP group (=7), a MT group (=7), a BNA+EA group (=8) and a shame BNA group (=7). Except of the sham BCP group, cancer cell inoculation operation at left tibia was given in the other 4 groups to establish the bone cancer pain model. In the MT group, the BNA+EA group and the shame BNA group, intraperitoneal injection of morphine hydrochloride was given to establish the morphine tolerance model. After the operation, bone-nearby acupuncture combined with electroacupuncture was applied at "Zusanli" (ST 36) and "Kunlun" (BL 60) in the BNA+EA group, with dilatational wave, 2 Hz/100 Hz in frequency, 0.5 to 1.5 mA in intensity. Intervention in the shame BNA group was applied at the same time and acupoints as those in the BNA+EA group, the needles were pierced the skin without any electrical stimulation. The needles were retained for 30 min, once a day for continuous 7 days in both BNA+EA and shame BNA groups. Before and 10, 11, 15, 22 days after the operation, the left paw withdrawal threshold (PWT) was measured in the 5 groups. The levels of HDAC1, HDAC2 and MOR in DRG were detected by Western blot.@*RESULTS@#Ten days after the cancer cell inoculation operation, the PWT of the BCP, MT, BNA+EA and sham BNA groups was decreased compared with the sham BCP group (0.05); the PWT of the BNA+EA group was increased compared with the MT and sham BNA group (<0.01). In the BCP group, the DRG levels of HDAC1 and HDCA2 were increased, while the level of MOR was decreased compared with the sham BCP group (<0.05, <0.01). In the MT group, the DRG level of HDAC1 was increased compared with the BCP group (<0.05). In the BNA+EA group, the DRG level of HDAC1 was decreased compared with the MT group and the sham BNA group (<0.01, <0.05), while the level of MOR was increased (<0.01).@*CONCLUSION@#Early intervention of bone-nearby acupuncture combined with electroacupuncture can relieve the morphine tolerance in bone cancer pain rats, it may relate to down-regulating the expression of HDAC1 and up-regulating the expression of MOR in the dorsal root ganglia.

Effect of the activation of P2X7 receptor on D-serine release in ventrolateral periaqueductal gray in rats with bone cancer pain / 医学研究生学报

Objective　The activation of P2X7 receptor in ventrolateral periaqueductal gray (vlPAG) is involved in the formation and maintenance of bone cancer pain (BCP). This study will establish a rat model of BCP and observe the effect of the activation of P2X7 receptor in vlPAG on D-serine level through brain microdialysis combined with ELISA.　Methods　Forty-two female SD rats were divided into four groups by random number table: normal control group (n=12), sham group (n=12), BCP group (n=12) and P2X7 receptor antagonist group (n= 6). The model of metastatic BCP in the tibias of the rats was established in the BCP group, and 20μL of RPMI-1640 medium cell suspension containing SHZ-88 breast cancer cells was injected (1×107 cancer cells/0.5 mL). The sham group was injected with treated cancer cells of the same volume (SHZ-88 breast cancer cells were kept in boiling water at 90 ℃ for 20 min), and the rest of the operation was the same as the BCP group. The normal control group received no treatment. The P2X7 receptor antagonist group was treated the same as the BCP group, except that the P2X7 receptor-specific antagonist A-438079 was added to the perfusion solution. The thermal pain threshold and mechanical pain threshold were detected at the same time in the normal control group, the sham group and the BCP group. The positive expression of P2X7 receptor in vlPAG of rats was detected by immunohistochemistry in each group in 21 days. The changes of D-serine in vlPAG dialysate were detected by ELISA in each group.　Results　The mechanical pain threshold and thermal pain threshold of the rats in BCP group on Day 5, 7, 10, 14, 18 and 21 were lower than those of the normal control group and sham group (P<0.01). The positive expression of P2X7 was scattered in vlPAG in normal control group and sham group. The number of P2X7 receptor positive cells in the BCP group was significantly higher than that in the control group and sham group (P<0.01). The content of D-serine in vlPAG of the rats in BCP group [(220.28±63.38)ng/mL] was significantly higher than that in the control group [(148.09±46.89)ng/mL] and the sham group [(147.32±51.44)ng/mL] (P<0.05). The content of D-serine in vlPAG [(134.20±41.77)ng/mL] in P2X7 receptor antagonist group was significantly lower than that in BCP group (P<0.05).　Conclusion　The activation of the P2X7 receptor in ventrolateral periaqueductal gray promotes D-serine release and participates in the mechanisms of BCP in rats .

Incidence and mortality of bone cancer among children, adolescents and young adults of Brazil

OBJECTIVES: Bone cancers occur frequently in children, adolescents, and young adults aging 15 to 29 years. Osteosarcoma and Ewing sarcoma are the most frequent subtypes in this population. The aim of this study was to describe incidence and mortality trends of bone cancers among Brazilian children, adolescents and young adults. METHODS: Incidence information was obtained from 23 population-based cancer registries. Mortality data were extracted from the Atlas of Cancer Mortality from 1979 to 2013. Specific and adjusted rates per million were analyzed according to gender, morphology and age at diagnosis. Median rates were used as a measure of central tendency. Joinpoint regression was applied to analyze trends. RESULTS: Median incidence rates were 5.74 and 11.25 cases per million in children and young adults respectively. Osteosarcoma in the 15-19 years aged group had the highest incidence rates. Stable incidence rates were observed among five registries in 0-14 year's age group. Four registries had a decreased incidence trend among adolescents and young adults. Median mortality rates were 1.22 and 5.07 deaths per million in children and young adults respectively. Increased mortality was observed on the North and Northeast regions. Decreased mortality trends were seen in the South (children) and Southeast (adolescents and young adults). CONCLUSION: Osteosarcoma and Ewing Sarcoma are the most incident bone cancers in all Brazilian regions. Bone cancers showed incidence and mortality patterns variation within the geographic regions and across age groups, although not significant. Despite limitations, it is crucial to monitor cancer epidemiology trends across geographic Brazilian regions.

Electroacupuncture intervention relieves pain possibly by promoting MOR endocytosis in locus coeruleus in bone cancer pain rats with morphine tolerance / 针刺研究

OBJECTIVE: To observe the effect of electroacupuncture (EA) on pain behavior and expression of µ-opioid receptor (MOR) and Rab5 (an important protein molecule for internalization of MOR) in the locus coeruleus (LC) region in bone cancer pain (BCP) rats with morphine tolerance (MT), so as to explore its mechanisms underlying improvement of BCP and MT. METHODS: The present study included two parts. In the first part, 23 female SD rats were randomized into sham BCP (n＝6), BCP (n＝9) and BCP＋MT (n＝8) groups, and in the second part, 61 female SD rats were randomized into 5 groups: sham BCP (n＝11), BCP (n＝11), BCP＋MT (n＝13), BCP＋MT＋EA (n＝13) and BCP＋MT＋sham EA (n＝13). The BCP morphine tolerance (BCP＋MT) model was established by injection of 10 µL of human Walker 256 breast cancer cells (MRMT-1 breast cancer cells, 1 x104 cells/µL) into the bone marrow cavity at the upper part of the left tibia and intraperitoneal injection of morphine hydrochloride (10 mg/kg, once per 12 h, for 11 successive days). On day 21 after inoculation, EA (2 Hz/100 Hz, 0.5－1.5 mA, increasing 0.5 mA every 10 min) was began to applied to bilateral "Zusanli" (ST30) and "Kunlun" (BL60) immediately after the first intraperitoneal injection of morphine. The treatment was performed for 30 min every time, once daily for 7 successive days. The paw withdrawal threshold (PWT) was detected before and 10, 11, 21, 22, 24, 26 and 28 days after inoculation. The immunoactivity of MOR and Rab5 proteins in the LC region was detected by immunofluorescence histochemistry. RESULTS: In the first part of the study, at the 10th day after inoculation of cancer cells, the PWT of the BCP and BCP＋MT groups was significantly lower than that of the sham BCP group (P0.05) but significantly lower than that of the sham BCP group (P0.05). CONCLUSION: EA intervention can relieve pain and MT in bone cancer pain rats with MT, which may be related to its effects in increasing MOR expression and promoting endocytosis of MOR in LC region.

Experimental study of metformin in relieving bone cancer pain by inhibiting JNK and improving gap junction function / 中国药科大学学报

@#This study aims to explore the involvement of c-Jun N-terminal kinase(JNK)-Gap junction regulation in the rat model of bone cancer pain and figure out whether adenosine 5′-monophosphate(AMP)-activated protein kinase(AMPK )activator metformin could attenuate bone cancer pain through this mechanism. Tumor cell implantation(TCI)induced bone cancer pain model in rats was established. The rats were administered, respectively, with 20 μL of metformin(50, 100 μg), JNK inhibitor SP600125(10 μg), gap junction inhibitor(carbenoxolone, CBX)(10 μg)and AMPK inhibitor Compound C(CC)(10 μg). The Von Frey Assay was applied to test the mechanical pain threshold. The activity of Glial fibrillary acidic protein(GFAP), ionized calcium-binding adaptor molecule 1(IBA-1)and Connexin 43(Cx43)in spinal cord was evaluated by immunohistochemistry. Changes of p-JNK expression were detected by Western blot. JNK inhibitor SP600125 relieved TCI-induced bone cancer pain significantly in rats, while this analgesic effect was almost canceled by the blocker of gap junction carbenoxolone(CBX). Various concentration of metformin(50, 100 μg, i. t. )significantly inhibited TCI-induced mechanical allodynia and the changes of p-JNK and p-Cx43 expression were also reversed in spinal cord in rats. Together, these data suggested that activation of AMPK with metformin attenuated TCI-induced bone cancer pain via regulating the function of JNK-Gap junction in rats.

Effects of CC chemokine ligand 2 on pain behavior in a rat model of bone cancer pain and its underlying peripheral mechanism / 上海交通大学学报（医学版）

Objective • To investigate the effects of CC chemokine ligand 2 (CCL2) on pain behavior in a rat model of bone cancer pain (BCP) and the underlying peripheral mechanisms. Methods • BCP models were developed by inoculation of Walker256 mammary gland carcinoma cells into the tibia medullary cavities of right hind limbs SD rats. The same volume of saline was injected in sham operation (sham) group. The mechanical pain threshold was measured to judge the success of BCP model. Expression of CCL2 in L4 and L5 dorsal root ganglion (DRG) was detected by immunofluorescence staining. CCL2 (500 ng, 25 μL) was injected into plantar of the operated side to observe its effects on leg-raising and foot-licking behaviors of hind paws in BCP and sham rats. Whole-cell patch-clamp recording was used to investigate the effects of CCL2 on membrane potential of acutely dissociated DRG neurons from the two groups. Results • Fourteen days after operation, the mechanical pain threshold in the right hind paws of BCP rats was significantly lower than that in sham rats. Compared with the Sham rats, the expression of CCL2 in L4 and L5 DRG of BCP rats was significantly higher. Plantar injection of CCL2 increased paw lift time in BCP rats. The rate and amplitude of depolarization induced by CCL2 in BCP DRG neurons were significantly higher than those in sham neurons. Conclusion • CCL2 facilitates pain behavior in BCP rats, and its peripheral mechanism maybe involves CCL2-induced neuron depolarization to enhance excitability of DRG neurons. These results indicate that CCL2 plays an important role in development of BCP.

Mechanism of spinal chemokine C-C motif receptor 2-mediated maintenance of bone cancer pain in rats / 上海交通大学学报（医学版）

Objective • To investigate the mechanism of spinal chemokine C-C motif receptor 2 (CCR2)-mediated maintenance of bone cancer pain (BCP) in rats. Methods • Fifty-four SD rats were divided into BCP group, sham operation group, BCP+INCB3344 (CCR2 specific antagonist) group, and BCP+vehicle control group. Walker256 breast cancer cells were injected into the tibia medullary cavity of rats in the BCP group to establish the BCP model, while the rats in the sham operation group were injected with the same amount of saline. The rats in the BCP+INCB3344 group received intrathecal injection of INCB3344 on the 14th day after the establishment of BCP model, while the BCP+vehicle control group rats were injected with the same amount of vehicle. The mechanical pain thresholds of BCP group rats and sham operation group rats were measured to judge the success of BCP model. The expressions of CCR2 in the dorsal horn of spinal cord in the sham operation group rats and the BCP group rats were detected by Western blotting. The effects of intrathecal administration of INCB3344 on the mechanical pain threshold of BCP rats were observed by mechanical pain behavior test. Whole-cell patch-clamp recordings were used to investigate the differences of spontaneous excitatory postsynaptic currents (sEPSCs), α-amino-3-hydroxy-5-methyl-4-isoxazole-propionicacid (AMPA) and N-methyl-D-aspartic acid (NMDA)-induced currents of spinal substantia gelatinosa (SG) neurons of rats in the BCP group, the BCP+INCB3344 group and the BCP+vehicle control group. Results • Compared with the sham operation group, the mechanical pain threshold of BCP group rats reduced significantly on the 14th day after operation (P=0.000), and the expression of CCR2 in ipsilateral spinal cord of BCP group rats increased significantly (P=0.009). After intrathecal injection of INCB3344 for 4 h, the mechanical pain threshold of BCP+INCB3344 group rats was significantly increased (P=0.002). The frequency and amplitude of sEPSCs and the amplitude of AMPA and NMDA-induced currents in SG neurons of BCP group rats were significantly higher than those of the sham operation group rats (all P=0.000), while intrathecal administration of INCB3344 could significantly inhibit the above-mentioned indices in the BCP+INCB3344 group (all P<0.05). In addition, extracellular perfusion of INCB3344 could also significantly inhibit the frequency (P=0.001) and amplitude (P=0.020) of sEPSCs in SG neurons in BCP rats. Conclusion • CCR2 expressing in the spinal cord mediates the enhancement of excitatory synaptic transmission efficacy in the spinal dorsal horn of BCP rats by enhancing the functions of AMPA and NMDA receptors, which may be an important mechanism for the maintenance of BCP.

Up-regulated expression of CD147 gene in malignant bone tumor and the possible induction mechanism during osteoclast formation

It is increasingly evident that the microenvironment of bone can influence cancer phenotype in many ways that favor growth in bone. CD147, a transmembrane protein of the immunoglobulin (Ig) superfamily, was identified independently in different species and has many designations across different species. However, expression levels of CD147 mRNA in bone cancer have not been described. In this study, we have used real-time fluorescence quantification (RT-PCR) to demonstrate CD147 expression in malignant bone cancer and benign bone tumor tissues. The results suggested that the expression of CD147 gene was significantly up-regulated in malignant bone cancer. Moreover, we found that over-expressed RANKL progressively enhanced osteoclast formation up to 48 h, which suggested that RANKL could promote the formation of osteoclast, indicating that both CD147 and RANKL play important roles in the formation of osteoclasts. Furthermore, the expressions of four osteoclast specific expression genes, including TRACP, MMP-2, MMP-9 and c-Src, were analyzed using RT-PCR. The results indicated that four osteoclast-specific expression genes were detectable in all osteoclast with different treatments. However, the highest expression level of these four osteoclast-specific expression genes appears in the CD147+ RANKL group and the lowest expression level of these four osteoclast-specific expression genes appears with si-RANKL treatment. Characterization of the role of CD147 in the development of tumors should lead to a better understanding of the changes occurring at the molecular level during the development and progression of primary human bone cancer.

Expression of IGF-1 in serum and phosphorylated IGF-1 receptor in spinal cord in mouse model of bone cancer pain / 中华行为医学与脑科学杂志

Objective To investigate the expression of insulin-like growth factors-1(IGF-1)in ser-um and phosphorylated IGF-1 receptor in spinal cord in mouse model of bone cancer pain. Methods Sixty male C3H/HeJ mice weighed 18-22 g were randomly divided into Sham group(n=30)and Tumor group(n=30). The mice in Tumor group were inoculated with NCTC fibrosarcoma cells in the right femur bone marrow cavity. Paw withdrawl mechanical threshold(PWMT)and the number of spontaneous flinches(NSF)were measured on 1d before inoculation and on 4 d,7 d,10 d,14 d,21 d after inoculation(n=8). At each time point,the mice of each group were taken blood by removal eyeball and the samples of blood were obtained to detect the expression of IGF-1 by enzyme-linked immunosorbent assay(n=4). The mice after taken blood on 14 d after inoculation were perfused and the samples of spinal cord lumber(L3~5)segment were obtained to detect the expression of phosphorylated IGF-1 receptor by immunofluorescence assay(n=6). Results Com-pared with Sham group,PWMT was significantly decreased(P<0.05)and NSF was significantly increased(P<0.05)on 7~21 d after inoculation. Compared with baseline value and Sham group(baseline value(27.33± 0.52)pg/ml,Sham group(29.11±1.86)pg/ml,(24.51±3.61)pg/ml,(23.33±4.59)pg/ml,(25.29±2.99) pg/ml),the expression of IGF-1 in serum was significantly increased on 7~21 d after inoculation in Tumor group((39.76±3.92)pg/ml,(36.93±2.18)pg/ml,(38.85±2.40)pg/ml,(39.70±2.62)pg/ml). The mean fluorescence intensity of phosphorylated IGF-1 receptor was significantly higher on 14d after inoculation in Tumor group(2.40±0.11)compared with Sham group(0.05±0.01). Conclusion Expression of IGF-1 in serum and phosphorylated IGF-1 receptor in spinal cord were significantly increased in mice with bone cancer pain,and this change may be involved in the development and maintenance of bone cancer pain.

Role of spinal cord CXCL13 in formation of bone cancer pain in rats / 重庆医学

Objective To investigate the role of spinal cord chemokine CXC ligand13(CXCL13) in the formation of rat bone cancer pain(BCP).Methods Twenty healthy female SD rats weighing 160-200 g were divided into four groups(n=5):sham operation group(S),BCP group(BP),small interference RNA(siRNA) negative control(NC-siRNA) group (NC) and CXCL13-siRNA group(CS).Normal saline was given by tibial medullary cavity injection in the S group.The tibial BCP model was established by tibial medullary cavity injection of equivalent Walker-256 breast cancer cells in the group BP,NC and CS.NC-siRNA lentivirus and CXCL13-siRNA lentivirus were injected intrathecally in the group NC and CS respectively.The mechanical pain threshold was measured on 1 d before model construction and on postoperative 7,9,14,21 d.The rats were killed after pain threshold measurement.The spinal cord and tibial tissue were taken.The co-expression of spinal CXCL13,microglia specific marker Iba-1 and neuron specific neucleoprotein NeuN was determined by using the immunofluorescence double standard staining,and expressions of CXCL13 and ionized calcium binding adaptor molecule-1 (Iba-1) protein and mRNA in spinal cord were detected by Western blot and RT-PCR;the HE staining microscopy was adopted to observe the tibial bone structure destroy situation.Results Compared with group S,the mechanical pain threshold in theBP group and NC group was decreased on 7-21 d after inoculation,CXCL13 expression in neuron was significantly increased and microglia was obviously activated,the expression of CXCL13 and Iba-1 protein and mRNA was significantly elevated (P＜0.05);compared with the NC group,the mechanical pain threshold on 9-21 d after model construction in the CS group was significantly increased,CXCL13 expression in neurons was significantly decreased,microglia activation was decreased and expression of CXCL13 and Iba-1 protein and mRNA was significantly decreased(P＜0.05);HE staining showed that the model groups appeared the tumor growth in bone marrow cavity,moreover which was eroded outwards and destroyed bone cortex,but no abnormality was found in the S group.Conclusion Spinal cord CXCL13 is involved in the BCP formation in rats by activating microglia.

Expression of spinal PD-L1 in bone cancer pain model mice and its effect on pain behaviors / 中华行为医学与脑科学杂志

Objective To evaluate the role of programmed cell death ligand-1 (PD-L1) in a mouse model of bone cancer pain.Methods Ninety-six male C3H/HeN mice (20-25 g,4-6 weeks old),which inoculated with osteolytic NCTC 2472 cells,were used to build the model of bone cancer pain.Part one:sixtyfour male C3H/HeJ mice were randomly divided into sham group (group Sham,n =32) and tumor group (group Tumor,n=32).Part two:Twenty-four male C3H/HeJ mice which were inoculated with osteolytic NCTC 2472 cells were randomly divided into group T (tumor,n=8),group PD-L1 (intrathecal injection with PLX3397,1 μg/5μl,n=8) and group NS (intrathecal injection with normal saline,n=8).Also,there were eight male C3H/HeJ mice in group S which were intra-femur inoculated with α-MEM.The pain behaviors of Sham group and Tumor group were observed and the expression of PD-L1 was detected before inoculation and on 4,7,10,14 and 21 days after inoculation,including paw withdrawal mechanical threshold (PWMT) and the number of spontaneous flinches (NSF).On 14 d after inoculation,the mice of group PD-L1 and group NS were intrathecal injected with drugs respectively.Pain behaviors were observed before injection and 2,4,6,24h after injection.Results Compared with group Sham,PWMT was significantly decreased and NSF was increased on 7～ 21 d after inoculation in group Tumor (P＜0.05).Compared with baseline and group S (baseline (0.38±0.06),group Sham (0.35±0.08),(0.38±0.08),(0.36±0.07)),the expression of PDL1 was up-regulated on 10-21 d after inoculation in group Tumor ((0.77±0.06),(1.21±0.04),(1.18±0.06)) (P＜0.05).Compared with group NS,PWMT was significantly increased (group NS (0.25t0.12),(0.25±0.12),(0.31±0.12),group PD-L1 (1.43±0.49),(1.35±0.44),(0.95±0.26)),and NSF was decreased on 2-6 h after injection in group PD-L1 (group NS(11.74± 1.31),(13.78±0.0.91),(13.63±1.06),group P D-L1 (4.90± 0.82),(4.15± 0.71),(7.65±0.56)) (P＜0.05).Conclusion Expression of PD-L1 in spinal cord was up-regulated in the mouse model of bone cancer pain.Intrathecal injection of recombinant PD-L1 has an analgesic effect on mice with bone cancer.

Osteochondroma in a young cat infected by feline leukemia virus / Ostecondroma em um felino jovem infectado pelo vírus da leucemia felina

ABSTRACT: Osteochondromas are primary bone tumors characterized by cartilage-covered bone projections involving single or multiple masses (osteochondromatosis). This study reports the clinical and pathological findings from a young domestic cat with osteochondroma in the humerus. During the clinical evaluation, the animal had pronounced right forelimb musculature atrophy and an increased distal humeral volume. Histopathological examination of the neoplasm revealed a proliferative lesion characterized mostly by endochondral ossification and peripheral foci of proliferating cartilage tissue. Further testing using immunohistochemical staining and polymerase chain reaction revealed the presence of feline leukemia virus antigens in the hematopoietic cells of the bone marrow and FeLV proviral DNA in the peripheral blood lymphocytes. Clinical and pathological findings are consistent with osteochondroma. This neoplasm occurred in an eight-month-old feline with humeral enlargement that had been present since two months old.

RESUMO: O osteocondroma é um tumor ósseo primário que se caracteriza por apresentar uma projeção óssea recoberta por cartilagem que pode ser formado por uma única ou por múltiplas massas (osteocondromatose). O objetivo deste trabalho é descrever os achados clínicos e patológicos de um felino jovem com osteocondroma no úmero. Na avaliação clínica, o felino apresentava atrofia acentuada da musculatura do membro torácico direito além de um aumento de volume em região distal do úmero. No exame histopatológico, observou-se uma lesão proliferativa composta em sua maior parte por ossificação endocondral com focos periféricos de tecido cartilaginoso proliferado. Testes de imuno-histoquímica e reação em cadeia da polimerase revelaram a presença de antígenos de FeLV em células da linhagem hematopoiética da medula óssea e DNA proviral em linfócitos do sangue periférico, respectivamente. Os achados clínicos e patológicos foram consistentes com um osteocondroma em um felino jovem, com oito meses de idade, com lesão em úmero desde os dois meses de idade.

Effect of intrathecal injection of TRPV3-siRNA lentivirus on pain behavior in rats with bone cancer pain / 中华行为医学与脑科学杂志

ObjectiveTo investigate the effect of intrathecal injection of TRPV3-siRNA lentivirus on bone cancer pain(BCP) behaviors in rats.Methods40 female SD rats successfully received intrathecal catheter implantation and without motor dysfunction were randomly divided into 4 groups (n=10 in each group):Sham group (S),BCP group (B),negative control lentivirus group (C) and TRPV3-siRNA lentivirus group (T).Group B,C and T were induced bone cancer pain by intra-right-femur inoculation of Wallker 256 cells,while rats in group S were injected of inactivated cell.Rats in group T were intrathecally treated with 5 μl TRPV3-siRNA lentivirus while rats in group C received 5 μl negative lentivirus on 1~6 d after surgery.All the rats received pain behaviors including paw withdrawal thermal latency(PWTL) and paw withdrawal mechanical threshold (PWMT) at 1 d before BCP and 1,3,6,9,12,15,18 and 21 d after BCP.L4~L6 spinal cords were reserved for RT-PCR and Western Blot.ResultsCompared with group S,PWTL and PWMT of group B were decreased (P0.05).The results of RT-PCR and Western blot demonstrated that the expression of TPPV3 in group T was decreased compared with that in group C(P<0.05).ConclusionIntrathecal injection of TRPV3-siRNA lentivirus can inhibit the expression of TRPV3 and thus alleviate symptom of PWTL,but not PWMT.

Research on the Inhibition of Breast Cancer Bone Metastasis Induced Rat Pain-related Behaviors by Intrathecally Injection of GSK690693, an Akt Specific Inhibitor / 现代生物医学进展

Objective:To investigate the role of Akt signaling pathway in the regulation of breast cancer bone metastasis induced pain behavior in rat.Methods:Model rats were randomly divided into three groups:Model group,Model+GSK690693 group and Model+ Saline group.On PID 13,14 and 21,Model+GSK690693 group rats were intrathecally injected with GSK690693,a specific inhibitor of Akt.Model+Saline group were injected with saline instead.The pain related behaviors were respectively recorded on PID 0,7,14 and 21.The expression ofp-Akt in DRG used for western bloting were examed on PID 21.Results:After the injection of Akt specific inhibitor GSK690693 by intrathecal,In the Model+GSK690693 group,the threshold value of mechanical contraction reflex was increased,the spontaneous pain behavior and the expression of p-Akt in DRG decreased.On PID 14 d and 21 d,the pain behavior of rats in Model+GSK690693 group was significantly different from that of Model+Saline group and Model group (P＜0.01);On PID 21 d,There was significant statistical significance (P＜0.01) on the expression of p-Akt and Model in the ipsilateral Model+GSK690693 of DRG group,which was statistically significant (P＜0.05) compared with the Model+Saline group.Conclusion:Intrathecal injection of Akt specific inhibitor GSK690693 inhibits rat pain related behaviors induced by bone metastasis in rat breast cancer.

Effects of electroacupuncture on expression of μ-opioid receptor in nucleus ceruleus in rats with bone cancer pain-morphine tolerance / 中国针灸

<p><b>OBJECTIVE</b>To observe the effects of electroacupuncture (EA) on pain behavior in rats with bone cancer pain and morphine tolerance, and to explore partial action mechanism.</p><p><b>METHODS</b>Forty-two SD healthy female rats were randomly divided into a sham operation group (7 rats), a bone cancer pain group (8 rats), a morphine tolerance group (9 rats), an EA group (9 rats) and a sham EA group (9 rats). The rats in the sham operation group were treated with injection of phosphate buffer saline at medullary cavity of left-side tibia, and the rats in the remaining groups were injected with MRMT-1 breast cancer cells. After operation, no treatment was given to rats in the sham operation group and bone cancer pain group. 11 days after operation, rats in the morphine tole-rance group, EA group and sham EA group were treated with intraperitoneal injection of morphine hydrochloride, once every 12 hours, for 11 days to establish the model of bone cancer pain and morphine tolerance. One day after the establishment of this bone cancer pain model, the rats in the morphine tolerance group were injected with morphine, once every 12 hours (9:00 a.m. and 9:00 p.m.) for 7 days; the rats in the EA group and sham EA group were injected with morphine at 9:00 a.m., and treated with EA (2 Hz/100 Hz) and sham EA (only injected into the subcutaneous tissue) at bilateral "Zusanli" (ST 36) and "Kunlun" (BL 60), 30 min per treatment, once a day for 7 days. One day before cancer cell injection, 6 days, 8 days, 10 days after operation, after 30 min on 1 days, 5 days, 9 days, 11 days of morphine injection, and after 30 min on 1 days, 3 days, 5 days, 7 days of EA treatment, the paw withdrawal threshold (PWT) was measured in each group. On 11 day of morphine injection, HE staining was applied to observe the morphology and structure change of tibia in the sham operation group, bone cancer pain group and morphine tolerance group, random 2 rats in each group. On 7 days of EA treatment, fluorescent immunohistochemical method was applied to observe the expression of μ-opioid receptor positive cells in nucleus ceruleus in each group, random 4 rats in each one.</p><p><b>RESULTS</b>After 10 days of the cancer cells injection, the PWT of 28 rats of bone cancer pain model (8 rats in the bone cancer pain group, 8 rats in the morphine tolerance group, 6 rats in the EA group and 6 rats in the sham EA group) was significantly lower than that of 7 rats in the sham operation group (<0.01). After one day of morphine injection, the PWT of the morphine tolerance group, EA group and sham EA group was higher than that of the bone cancer pain group (all<0.01); on 11 d of morphine injection, the PWT of the morphine tolerance group, EA group and sham EA group was not significantly different from that of the bone cancer pain group (all>0.05). On 11 d of morphine injection, the tumor induced by cancer cells was observed in upper 1/3 tibia in the bone cancer pain group and morphine tolerance group, and the marrow cavity was filled with MRMT-1 cancer cells; no abnormal change was observed in the sham operation group. On 1 d, 3 d, 5 d and 7 d of EA treatment, the PWT of the cancer pain group, morphine tolerance group and sham EA group was lower than that of the EA group (all<0.01). On 7 d of EA treatment, the positive expression of MOR in nucleus ceruleus in the cancer pain group, morphine tolerance group, EA group and sham EA group was lower than that in the sham operation group (<0.01,<0.05), and that in the cancer pain group, morphine tolerance group and sham EA group was lower than that in the EA group (all<0.01).</p><p><b>CONCLUSIONS</b>EA can improve mechanical pain threshold in rats with bone cancer pain-morphine tolerance, and improve the abnormal pain, which is likely to be involved with improvement of the MOR positive cells expression in nucleus ceruleus by EA.</p>

Effect of butorphanol and sufentanil with epidural injections on neurobehavior function in bone cancer pain model rats / 中华行为医学与脑科学杂志

Objective To observe the effect of different doses of butorphanol and sufentanil repeatedly epidural injected on the neurobehavior function in bone cancer model rats.Methods A PE-530 catheter was inserted into the epidural space of all male Sprague-Dawley rats(not mated,weighting 150-180 g) at L1-2 level.Three days after operation,64 rats without any motor dysfunction were randomly divided into eight groups (n=8):sham operated group (group C),normal saline with bone cancer pain group (group N),butorphanol groups(group B 1,B2,B3)and sufentanil groups (group S1,S2,S3).Bone cancer pain model was constructed in group N,B and S when rats in group C were sham operated.Rats in group C and N were epidurally injected NS 30μl each,and rats in group B1,B2 and B3 were respectively epidurally injected butorphanol 25,50,100 μg (all diluted to 30 μl with NS),when rats in group S1,S2 and S3 were respectively cpidurally injected sufentanil 1,2,4 μg (all diluted to 30 μl with NS) on time per day for 10-14 days after modeling.The neurobehavior paw withdrawal threshold (MWT) of the left hind claw was recorded to observe the changes in pain behavior.The neurobehavior function of rats were recorded by BBB (BASSO,BEATTIE and BRESNAHAN) score and the inclined plane test.Results Compared with group C((67.65±9.29) g),the MWT of the model groups obviously decreased before the first time of injection (N (15.23± 2.46) g,B 1 (16.14±2.28) g,B2(15.42±3.22) g,B3(14.35±2.32) g,S1 (15.37±2.11)g,S2(15.22±2.93) g,S3(16.25± 2.36) g) (all P＜0.05)).Compared with group N((16.13±2.37) g),the MWT of group B2,B3 and S3 increased obviously after the first time of injection ((35.12±5.16) g,(35.63± 1.53) g and (34.24±5.93) g) (P＜ 0.05).At the first day of injection,there was no significant difference in the BBB scores and the inclined plane test between the model groups (P＞0.05).At 6 h after the forth injection the inclined plane test and the BBB scores of group B3 were obviously decreased compared with group N ((34.72 ± 4.56) ° and (10.64 ± 1.82) points to (43.15±4.67)° and (14.05±1.78) points (P＜0.05)).Conclusion The results provide evidence that repeatedly epidural injection of butorphanol 50 μg or 100 μg or sufentanil 4 μg can reduce the pain of the rats with bone cancer pain.But repeated epidural injection of butorphanol 100 μg can injure the neurological function.

Bone cancer pain induce anxiety-like behavior and high expression of NR2B subunit in anterior cingulate cortex of rats / 临床麻醉学杂志

Objective To investigate the effect of bone cancer pain on emotion and NMDA re-ceptor NR2B subunit expression level in anterior cingulate cortex (ACC)in rats.Methods One hun-dred and fifty healthy male Wistar rats weighing 200-250 g aged 3 months old were randomly divided into 3 groups (n = 50 in each group):sham operation group (group S),bone cancer pain group (group BCP),RO25-6981 group (group RO).The BCP model was established by inoculating Walker 256 breast cancer cells into right intra-tibial.Rats in group S were given the same dose of d-hanks. Group RO was injected intraperitoneally with RO25-6981 (5 mg/kg/d)on the day of inoculation, while rats in the group S and group BCP were given the same dose of normal saline.Mechanical with-drawal threshold (MWT)and thermal withdrawal latency (TWL)of right hind legs were measured on day 7,10,14 after inoculation respectively.Elevated plus-maze test was carried out to investigate the effect of bone cancer pain on emotion in rats after pain threshold detection,then the percentage of the times entering the open arms (OE)and the percentage of the time staying in the open arms (OT) duration the total period were evaluated.Then the anterior cingulate cortex tissue was removed to e-valuate the NR2B protein and mRNA expression levels by RT-PCR,Western blot and immunofluo-rescence methods on day 14 after elevated plus-maze test.Results All the parameters did not differ with significant difference between group S and group RO.MWT decreased and TWL shortened on day 7,10,14 after inoculation in group BCP compared with those before inoculation and those of group S and group RO.OE and OT in group BCP reduced remarkably than those before inoculation and those of group S and group RO.Relative absorbance of NR2B mRNA,the expression of NR2B pro-tein,average NR2B relative fluorescence intensity value is obviously higher than that of group S and group RO (P <0.05).Conclusion Bone cancer pain can induce pain-related aversion and anxiety-like behavior of rats,and the mechanism may be related to the high expression of NR2B in anterior cingu-late cortex.

JNK in spinal cord facilitates bone cancer pain in rats through modulation of CXCL1 / 华中科技大学学报（医学）（英德文版）

In patients with advanced cancer, cancer-induced bone pain (CIBP) is a severe and common problem that is difficult to manage and explain. As c-Jun N-terminal kinase (JNK) and chemokine (C-X-C motif) ligand 1 (CXCL1) have been shown to participate in several chronic pain processes, we investigated the role of JNK and CXCL1 in CIBP and the relationship between them. A rat bone cancer pain model was established by intramedullary injection of Walker 256 rat gland mammary carcinoma cells into the left tibia of Sprague-Dawley rats. As a result, intramedullary injection of Walker 256 carcinoma cells induced significant bone destruction and persistent pain. Both phosphorylated JNK1 (pJNK1) and pJNK2 showed time-dependent increases in the ipsilateral spinal cord from day 7 to day 18 after tumor injection. Inhibition of JNK activation by intrathecal administration of SP600125, a selective pJNK inhibitor, attenuated mechanical allodynia and heat hyperalgesia caused by tumor inoculation. Tumor cell inoculation also induced robust CXCL1 upregulation in the ipsilateral spinal cord on day 18 after tumor injection. Inhibition of CXCL1 by intrathecal administration of CXCL1 neutralizing antibody showed a stable analgesic effect. Intrathecal administration of SP600125 reduced CXCL1 increase in the spinal cord, whereas inhibition of CXCL1 in the spinal cord showed no influence on JNK activation. Taken together, these results suggested that JNK activation in spinal cord contributed to the maintenance of CIBP, which may act through modulation of CXCL1. Inhibition of the pJNK/CXCL1 pathway may provide a new choice for treatment of CIBP.

Change in expression of WNK1 in spinal cord of a rat model with bone cancer pain / 中国药理学通报

Aim To investigate the changes in the ex-pression of WNK1 in spinal cord of a rat model with bone cancer pain. Methods Female SD rats, weig-hing 170 ~200 g, were randomly divided into three groups:normal control group (group C, n=3), sham operation group ( group S, n =3 ) and bone cancer pain group ( group BCP, n =24 ) . Group C was not given any treatment, and group S was injected into the bone marrow of left tibia with 5 μl PBS solution while group BCP with 5 μl WALKER 256 mammary gland cancer cell suspension (approximately 1 × 105 cells). Mechanical paw withdrawal threshold ( MWT ) was measured at d1 before inoculation ( baseline) and d3, 6,9,10,11,12 after inoculation. Group S and C were sacrificed at d 12 while group BCP at d 3 ,6 ,9 ,12 after inoculation and spinal cord ( L4~6 ) were removed at different time points for detection of WNK1 mRNA ex-pression by qRT-PCR and WNK1 protein expression by Western blot. Results Compared with group C and S,group BCP’ s MWT started to decrease since d 3 ( P0. 05 ) while the protein expression upregulated since d6 and also showed an in-creasing trend to d 12 ( P<0. 01 ) . Conclusion The expression of WNK1 in spinal cord of a rat model with bone cancer pain increased abnormally, which may be involved in the occurrence and maintenance of a rat model with bone cancer pain.