Effect of Rolipram on in vitro maturation, gene expression and embryonic development in bovines / Efeito de rolipram na maturação in vitro, expressão gênica e desenvolvimento embrionário em bovinos

The aim of this work was to evaluate the effect of the Rolipram during the maturation of bovine oocytes and gene expression of embryos produced in vitro. Bovine ovaries were collected in slaughterhouse. The COCs were selected and divided into 5 groups: Control 0 time; Control: IVM for 24 hours; Rolipram treatments with IVM blocking for 24 hours in maturation medium containing (100, 150 and 200µM). After 24 hours all groups were reseated in IVM for another 24 hours. Subsequently COCs were subjected to the same IVM system and fertilized, being checked for cleavage post fertilization and for blastocyst. In addition, performed expression of the following genes: Mater, BMP15 and Bax. No difference was found in gene expression. Of oocytes evaluated shortly after follicular aspiration, 79.00% were in GV, GVBD, MI, while 13.40%, were in MII and 7.60%, D/NI. Significant difference was observed in different concentrations (T100, T200 and T150µM) in oocytes that have reached the MII phase compared to control treatments (P= 0.003). Differences were observed in cleavage rate (P< 0.05) between T150 and T200 when compared to the C/24 Group. A high difference was observed on blastocyst rate (P< 0.001) among treatments compared to the control group.(AU)

O objetivo deste trabalho foi avaliar o efeito do rolipram durante a maturação de oócitos bovinos, expressão gênica e embriões produzidos in vitro. Os ovários bovinos foram coletados no matadouro. Os COCs foram selecionados e divididos em cinco grupos: controle 0 tempo; controle: MIV por 24 horas; tratamentos rolipram com bloqueio MIV por 24 horas em meio de maturação contendo 100, 150 e 200µM. Após 24 horas, todos os grupos foram recolocados em MIV por mais 24 horas. Subsequentemente COCs foram submetidos ao mesmo sistema MIV e fertilizados, sendo avaliada a taxa de clivagem e de blastocisto, além da expressão dos seguintes genes: Mater, BMP15 e Bax. Nenhuma diferença foi observada na expressão gênica. Dos oócitos avaliados logo após a aspiração folicular, 79,0% estavam em GV, GVBD, MI, enquanto 13,40% estavam em MII, e 7,60% em D/NI. A diferença significativa foi observada em diferentes concentrações (T100, T200 e T150µM) em oócitos que atingiram a fase MII em comparação aos tratamentos de controle (P=0,3). Diferenças foram observadas nas taxas de clivagem (P<0,5) entre T150 e T200 quando comparadas com as taxas do grupo C/24. Uma grande diferença foi observada na taxa de blastocisto (P<0,1) entre os tratamentos em relação ao grupo controle.(AU)

Phosphodiesterase 4 inhibitor rolipram prevents chronic alcoholism and withdrawal-induced depression-like behaviors in mice / 中国病理生理杂志

AIM:To investigate the effect of phosphodiesterase 4 (PDE4) inhibitor rolipram on the levels of cyclic adenosine monophosphate (cAMP), protein kinase A (PKA), cAMP response element-binding protein (CREB), phosphorylated CREB (p-CREB) and brain-derived neurotrophic factor (BDNF) in the hippocampus and the prefrontal cortex (PFC) of alcoholism model mice.METHODS:The mice (n=60) were randomly divided into control group , con-trol+rolipram group, alcoholism model group, and alcohol +rolipram (0.1, 0.5 and 1 mg/kg) groups.The mice were given alcohol preference test on days 6, 13, 20 and 27.After the test, the mice received withdrawal of alcohol for 1 d.On day 28, the mice were given behavior test of depression , and after the test, the mice were sacrificed.The cAMP levels in the hippocampus and PFC were detected by ELISA , and the protein levels of PKA , CREB, p-CREB and BDNF were detec-ted by Western blot.RESULTS:The mice showed an obvious drinking phenomenon (P<0.01), and the immobility time of forced swimming test and tail suspension test was significantly increased (P<0.01), with increasing drinking days and withdrawal times .However , chronic treatment with rolipram for 28 d reversed this phenomenon .Moreover , the cAMP lev-els in the hippocampus and PFC were significantly decreased after 28 d alcohol treatment ( P<0.01 ) , and pretreatment with rolipram (1 mg/kg) obviously reversed this decrease (P<0.01).Parallel to these changes of cAMP , the protein lev-els of PKA, p-CREB and BDNF were also decreased in the hippocampus and PFC (P<0.01), and 28 d rolipram adminis-tration inhibited the decreased cAMP , PKA, p-CREB and BDNF levels in the hippocampus .Moreover, 28 d rolipram ad-ministration also reversed decreased cAMP , PKA and p-CREB in the PFC.CONCLUSION:Rolipram treatment protects against alcohol-induced depression-like behaviors , and also reduces alcohol drinking .These effects may be related to PDE4-cAMP-PKA-CREB-BDNF pathway .

Rolipram represses MRP8/14-induced pro-inflammatory cytokine releases in RAW264.7 cells through NF-κB activation / 中国病理生理杂志

AIM:To investigate the inhibitory effect of PDE4 inhibitor rolipram on the releases of inflammatory cytokines in mouse macrophages (RAW264.7 cells) induced by myeloid-related protein 8/14 (MRP8/14), and to explore the role of nuclear factor-κB (NF-κB) in this process.METHODS:RAW264.7 cells were treated with MRP8/14.The inflammatory cytokines TNF-α and IL-6 were determined by LiquiChip and qPCR.In contrast, RAW264.7 cells were pretreated with rolipram prior to MRP8/14 exposure.After MRP8/14 challenge, the inflammatory cytokines TNF-α and IL-6 were determined by LiquiChip and qPCR.Moreover, the phosphorylation level of NF-κB P65 was determined by Western blot.The nuclear translocation of NF-κB P65 in RAW264.7 cells was detected by indirect immunofluorescence.RESULTS:The results of LiquiChip and qPCR showed that MRP8/14 enhanced the expression of TNF-α and IL-6 (P<0.01),while pretreatment with rolipram markedly down-regulated the level of inflammatory cytokines induced by MRP8/14 (P<0.05).Meanwhile, MRP8/14 significantly activated the phosphorylation of NF-κB P65, and the protein expression of p65 in the nucleus was increased,while pretreatment with rolipram suppressed the phosphorylation of NF-κB P65 induced by MRP8/14.CONCLUSION:Rolipram may significantly reduce the releases of the inflammatory cytokines induced by MRP8/14 through the NF-κB signaling.

Protective Effect of Administered Rolipram against Radiation-Induced Testicular Injury in Mice

PURPOSE: Pelvic irradiation for the treatment of cancer can affect normal cells, such as the rapidly proliferating spermatogenic cells of the testis, leading to infertility, a common post-irradiation problem. The present study investigated the radioprotective effect of rolipram, a specific phosphodiesterase type-IV inhibitor known to increase the expression and phosphorylation of the cyclic adenosine monophosphate response element-binding protein (CREB), a key factor for spermatogenesis, with the testicular system against pelvic irradiation. MATERIALS AND METHODS: Male C57BL/6 mice were treated with pelvic irradiation (2 Gy) and rolipram, alone or in combination, and were sacrificed at 12 hours and 35 days after irradiation. RESULTS: Rolipram protected germ cells from radiation-induced apoptosis at 12 hours after irradiation and significantly increased testis weight compared with irradiation controls at 35 days. Rolipram also ameliorated radiation-induced testicular morphological changes, such as changes in seminiferous tubular diameter and epithelial height. Additionally, seminiferous tubule repopulation and stem cell survival indices were higher in the rolipram-treated group than in the radiation group. Moreover, rolipram treatment counteracted the radiation-mediated decrease in the sperm count and mobility in the epididymis. CONCLUSIONS: These protective effects of rolipram treatment prior to irradiation may be mediated by the increase in pCREB levels at 12 hours post-irradiation and the attenuated decrease in pCREB levels in the testis at 35 days post-irradiation in the rolipram-treated group. These findings suggest that activation of CREB signaling by rolipram treatment ameliorates the detrimental effects of acute irradiation on testicular dysfunction and the related male reproductive functions in mice.

Research progress of PDE4 in the central nervous system / 中国药理学通报

Phosphodiesterase-4 ( PDE4 ) has been one of the most popular drug targets during recent years due to its critical role in the control of intracellular cyclic adenosine monophos-phate ( cAMP ) concentration and downstream signal transduc-tion. PDE4 is widely distributed in the central nervous system ( CNS) with different expression levels of its four subtypes. Re-cent data indicate that altered PDE4 expression and/or activity is relevant to multiple CNS disorders, such as depression, memory deficiency, drug dependence, and neural lesion. Selective PDE4 inhibitors exhibit therapeutic effects on these disorders and might be a promising pharmacotherapy. The paper highlights recent re-search progress in the roles of PDE4 in CNS function, and dis-cusses the prospects of PDE4 as a novel therapeutic target for CNS disorders.

A potential novel antidepressant target based on phosphodiesterase 4: Research advances / 国际药学研究杂志

Depression is a prevalent and life-threatening mental disorder. Its mechanisms remain unclear. It was found that depressive disorders are closely related to monoamine neurotransmitters absence, down-regulation of hypothalamic-pituitary-adrenal (HPA) axis and neurotrophic factors and dendritic complexity in brain. Phosphodiesterase 4 (PDE4) inhibitor rolipram produce antidepressant and memory-enhancing effects via intracellular cAMP signaling. However, its clinical utility is limited by the major side effect of emesis, which appears to be PDE isoform-specific. Therefore, this research is to investigate whether long-form PDED variants mediate antidepressant and cognition-enhancing effects, without causing emesis. This review summaries the PDE molecular structure, distribution and its antidepressant function, so as to illustrate the advantage of selective subtype inhibitor in the treatment of depression.

Electrophysiological and Behavioral Changes by Phosphodiesterase 4 Inhibitor in a Rat Model of Alcoholic Neuropathy

OBJECTIVE: Alcoholic neuropathy is characterized by allodynia (a discomfort evoked by normally innocuous stimuli), hyperalgesia (an exaggerated pain in response to painful stimuli) and spontaneous burning pain. The aim of the present study is to investigate the effect of rolipram, a phosphodiesterase 4 inhibitor, against alcohol-induced neuropathy in rats. METHODS: Allodynia was induced by administering 35% v/v ethanol (10 g/kg; oral gavage) to Spraue-Dawley rats for 8 weeks. Rolipram and saline (vehicle) were administered intraperitoneally. Mechanical allodynia was measured by using von Frey filaments. Somatosensory evoked potential (SEP) was proposed as complementary measure to assess the integrity of nerve pathway. RESULTS: The ethanol-induced mechanical allodynia began to manifest from 3 week, and then peaked within 1 week. Beginning from 3 week, latency significantly started to increased in control group. In rolipram treated rats, the shorter latency was sustained until 8 weeks (p<0.05). The mechanical allodynia, which began to manifest on the 3 weeks, intraperitoneal injections of rolipram sustained statistical difference until 8 weeks, the final week of the study (p<0.05). CONCLUSION: This study suggests that rolipram might alleviate mechanical allodynia induced by alcohol in rats, which clearly has clinical implication.

The effects of rolipram on neurofunction and the ultrastructure of the spinal cord after spinal cord contusion / 中华物理医学与康复杂志

Objective To investigate the effects of rolipram on neurofunction and the ultrastructure of the spinal cord in rats with spinal cord contusion.Methods Thirty adult,female Sprague-Dawley (SD) rats received spinal cord contusion at the T10 level.They were then randomited into an experimental group and a control group immediately after the operation.Rats in the experimental group received subcutaneous injections of 0.25mg/kg of rolipram twice daily for two weeks.Control rats received the same dosage of 0.9％ sodium chloride solution on the same schedule.The rats'functional recovery was evaluated using the open-field locomotion rating scale of Basso,Beattie and Bresnahan (BBB score),once a week within the 1st month after spinal cord injury (SCI),and once every two weeks subsequently.The morphology of the spinal cord tissue around the lesion site was observed under the light microscope with HE staining at the 8th week postoperation,and the ultrastructure of the spinal cord was observed under the transmission electron microscope at the 2nd,4th and 8th week postoperation.Results At the 2nd and 3rd week after SCI the experimental group exhibited significandy greater improvement in average BBB scores than the control group.However,the average BBB scores in the experimental and control groups were not significantly different at 8 weeks after SCI.Under the light microscope,cavities were observed in the posterior dorsal column near the SCI in both the experimental and control groups.However thick,condensed glial scars in the injured area were observed only in the control group.The density of glial cells decreased more in the experimental group than in the control group.Transmission electron microscopy revealed that,compared with the control group,inflammatory edema was attenuated and fewer axons were damaged at the 2nd week postoperation in the experimental group.That group also showed less axon degeneration as well as more angiogenesis at the 4th and 8th week.Conclusion Rolipram can reduce secondary neuron degeneration,inhibit gliosis and the formation of glial scars,and promote angiogenesis.

The Effect of Phosphodiesterase-4-Specific Inhibitor in the Rat Model of Spinal Nerve Ligation

OBJECTIVE: Peripheral neuropathy is characterized by hyperalgesia, spontaneous burning pain, and allodynia. The purpose of this study was to investigate the effect of rolipram, a phosphodiesterase-4-specific inhibitor, in a segmental spinal nerve ligation model in rats. METHODS: Both the L5 and L6 spinal nerves of the left side of the rats were ligated. Phosphodiesterase-4 inhibitor (rolipram) and saline (vehicle) were administered intraperitoneally. We measured mechanical allodynia using von Frey filaments and a nerve conduction study. RESULTS: The mechanical allodynia, which began to manifest on the first day, peaked within 2 days. Multiple intraperitoneal injections of rolipram ameliorated the mechanical allodynia. Furthermore, an intraperitoneal administration of rolipram improved the development of pain behavior and nerve conduction velocity. CONCLUSION: This study suggests that the phosphodiesterase-4 inhibitor, rolipram, alleviates mechanical allodynia induced by segmental spinal nerve ligation in rats. This finding may have clinical implications.

The Modulation of IL-12 by Phosphodiesterase IV Inhibitor and Subsequent Changes of Th-1, Th-2 Cytokines in Human Endometrial Tissue / 대한산부인과학회잡지

OBJECTIVE: To assess the capability of phosphodiesterase type IV inhibitor (rolipram) to suppress IL-12 in human decidua and the subsequent changes of Th-2 cytokine (IL-10) and Th-1 cytokine (TNF-alpha). METHODS: Decidual tissues of 10 first-trimester pregnant women and 10 first-trimester pregnant women diagnosed as missed abortion were collected by dilatation and currettage. The decidual tissues were treated with rolipram for 6 hours. Protein and mRNA expression in the tissues were analysed by western blotting, immunohistochemistry and reverse transcription-polymerase chain reaction. RESULTS: Rolipram, in the concentration above 1 microgram/ml, could decrease the expression of IL-12p35 (control: 46.37+/-7.38, rolipram: 24.34+/-8.46) and IL-12p40 mRNA (control: 31.7+/-5.8, rolipram: 14.9+/-4.6) and protein (control: 52.4+/-8.9, rolipram: 40.9+/-12.1). However, the expression of IL-10 and TNF-alpha mRNA and protein did not changed by rolipram. There was no difference in the cytokine expression pattern between the decidual tissues of normal pregnancy and missed abortion. CONCLUSION: Rolipram, the phosphodiesterase type IV inhibitor, could induce the decrease of IL-12 in human decidua. In human decidual tissue, unlike other human tissues, the decrease of IL-12 by rolipram did not modulate other Th-1/Th-2 cytokines. Inability of IL-12 to modulate other Th-1/Th-2 cytokines might be related with unique cytokine network in human decidua rather than its small extent of decrease.

Effects of rolipram on learning and memory and the activity of PDE4 in hippocampus following the focal brain injury induced by ischemia-reperfusion in rats / 中国病理生理杂志

AIM:To investigate the effects of rolipram on the ability of learning and memory and the activity of PDE4 in hippocampus following the focal brain injury induced by ischemia-reperfusion in rats.METHODS:The cerebral ischemia-reperfusion injury model was made by middle cerebral artery occlusion(MCAO) in rats.The rats were randomly divided into sham-operated group,model group,and rolipram group.Rolipram was administered once a day(1 mg/kg,ip) from 6 h after the onset of the operation for 2 weeks.Then the learning and memory abilities were tested after Morris water maze and step-though training.The activity of PDE4 in hippocampus was evaluated by HPLC.RESULTS:In the Morris water maze test,compared to sham-operated group,the platform-finding time and swimming distance in model group were significantly longer(P