Application Research of Individualized Conditional Reprogramming System to Guide Treatment of Gastric Cancer.

BACKGROUND: Gastric cancer (GC) is one of the most common causes of malignant tumors in the world. Due to the high heterogeneity of GC and lack of specificity of available chemotherapy regimens, these tumors are prone to resistance, recurrence, and metastasis. Here, we formulated an individualized chemotherapy regimen for GC using a modified individual conditional reprogramming (i-CR) system. We established a primary tumor cell bank of GC cells and completed drug screening in order to realize individualized and accurate GC treatment. METHODS: We collected specimens from 93 surgical or gastroscopy GC cases and established a primary tumor cell bank using the i-CR system and PDX models. We also completed in vitro culture and drug sensitivity screening of the GC cells using the i-CR system. Whole-exome sequencing (WES) of the i-CR cells was performed using P0 and P5. We then chose targeted chemotherapy drugs based on the i-CR system results. RESULTS: Of the 72 cases that were collected from surgical specimens, 26 cases were successfully cultured with i-CR system, and of the 21 cases collected from gastroscopy specimens, seven were successfully cultured. Among these, 20 cases of the PDX model were established. SRC ± G3 had the highest culture success rate. The i-CR cells of P0 and P5 appeared to be highly conserved. According to drug sensitivity screening, we examined the predictive value of responses of GC patients to chemotherapeutic agents, especially in neoadjuvant patients. CONCLUSION: The i-CR system does not only represent the growth characteristics of tumors in vivo, but also provides support for clinical drug use. Drug susceptibility results were relatively consistent with clinical efficacy.

Characterization of Age-dependent Behavior Deficits in the PGC-1α Knockout Mouse, in Relevance to the Parkinson's Disease Model.

Parkinson's disease is a disorder of adult onset involving the progressive degeneration of selective portions of the central nervous system. It is known that mitochondrial dysfunction is involved in the pathogenesis of PD. Given that PGC-1α induces proliferation of mitochondria via transcription regulation, it is possible that PGC-1α pathway dysregulation is involved in PD pathogenesis. To determine how derangement of the PGC-1α pathway in age contributes to PD, in this study, we have characterized the number of dopaminergic neuron in the substantia nigra pars compacta (SNpc), motor behaviors and related expression of mitochondrial markers (CoxIV, SDHA, and Tomm20) in the ventral midbrains of PGC-1α null mice. We found an overall decrease in spontaneous, voluntary movements and severely impaired motor coordination in all age groups (10 months and 20 months) of PGC-1α null mice, while pole testing detected impaired motor activity in older PGC-1α null mice only. TH-positive neurons were significantly less in older PGC-1α null mice. Concentration of DA as well as its two metabolites reduced in an age-dependent manner in PGC-1α null mice. Expression of CoxIV, SDHA and Tomm20 also significantly decreased in the ventral midbrains of 10-month-old PGC-1α null mice. Thus, PGC-1α KO in mice induced dopaminergic neuron degeneration in the SNpc and DA deficits in the striatum in an age-dependent manner. Progressive impairment of motor coordination in an age-dependent manner was correlated to the extent of nigrostriatal dopaminergic pathway degeneration and mitochondrial dysfunction.

Changes in the Morphology, Number, and Pathological Protein Levels of Plasma Exosomes May Help Diagnose Alzheimer's Disease.

Exosomes are nano-sized extracellular vesicles that are secreted by cells and usually found in body fluids. Since they freely cross the blood-brain barrier, neuronal exosomes respond directly to changes in the brain's environment. Recent studies have shown that exosomes contain both amyloid-ß (Aß) and tau proteins and have a controversial role in the Alzheimer's disease (AD) process. In this study, enzyme-linked immunosorbent assay was used to detect the levels of P-S396-tau and Aß1-42 in plasma exosomes. We found that levels of P-S396-tau and Aß1-42 in plasma exosomes of AD patients were significantly higher compared to those in matched healthy controls. The difference between plasma exosomes of AD patients and those of matched healthy controls was determined using transmission electron microscopy and nanoparticle tracking analysis. Exosomes from AD patients were smaller and lower in quantity. These data together may provide a basis for early diagnosis of AD.

Neuroprotective Effect of Resveratrol via Activation of Sirt1 Signaling in a Rat Model of Combined Diabetes and Alzheimer's Disease.

BACKGROUND: Alzheimer's disease (AD) and diabetes mellitus (DM) often coexist in patients because having one of these conditions increases risk for the other. These two diseases share several pathophysiological mechanisms, such as specific inflammatory signaling pathways, oxidative stress, and cell apoptosis. It is still unclear exactly which mechanisms associated with DM are responsible for increased AD risk. Studies have found that even transient elevation of brain Aß levels can allow T2DM to slightly disrupt the neural milieu in a way that encourages pathologies associated with the onset of memory deficits and AD. A recent study argues that a potential common pathogenetic mechanism underlying both DM and AD is evidenced by the cooccurrence of amyloid brain legions and deposits containing both tau and Aß in pancreatic ß cells. Given these links, an investigation detailing disease mechanisms as well as treatment options for patients with cooccurring DM and AD is urgently needed. The biological effects of resveratrol relevant to DM and AD treatment include its abilities to modulate oxidative stress and reduce inflammation. A rat model of DM and concomitant AD was created for this study using intraperitoneal injection of streptozotocin and hippocampal injection of Aß1-40 to characterize resveratrol's potential protective action. RESULTS: Resveratrol significantly increased the Sirt1 expression, inhibited the memory impairment, the increased acetylcholinesterase, malondialdehyde, interleukin-1ß and interleukin 6 levels, and the decreased levels of choline acetyltransferase (ChAT), superoxide dismutase (SOD), and glutathione in this rat model of diabetes and concomitant AD. The Sirt 1 inhibitor EX527 partially reversed the effects of resveratrol. CONCLUSION: This study suggests that resveratrol may have a neuroprotective action through activation of Sirt1 signaling in diabetes and AD with concurrent onset.

A Pilot Study of Urinary Exosomes in Alzheimer's Disease.

BACKGROUND: Exosomes are nano-sized extracellular vesicles secreted by most cell types and abundantly present in body fluids, including blood, saliva, urine, cerebrospinal fluid, and breast milk. Exosomes can spread toxic amyloid-beta (Aß) and hyperphosphorylated tau between cells, contributing to neuronal loss in Alzheimer's disease (AD). OBJECTIVE: To explore changes in the morphology, number, and pathological protein levels of urinary exosomes in AD patients compared with age-matched healthy subjects. METHODS: In this study, enzyme-linked immunosorbent assay was used to detect the levels of Aß1-42 and P-S396-tau (normalized by CD63) in urinary exosomes of AD patients and matched healthy subjects. We used transmission electron microscopy and nanoparticle tracking analysis to observe the exosomes. RESULTS: We found that the levels of Aß1-42 and P-S396-tau in the urinary exosomes of AD patients were higher than those of matched healthy controls. Exosomes taken from AD patients were more numerous. CONCLUSION: The differences in levels of Aß1-42 and P-S396-tau and the quantity of urinary exosomes between AD patients and healthy controls may provide a basis for early diagnosis of AD.

The role of MAPT gene in Chinese dementia patients: a P301L pedigree study and brief literature review.

BACKGROUND AND PURPOSE: Frontotemporal dementia (FTD) is the second most common presenile dementia characterized by behavioral changes and language impairment. The diagnosis of FTD relies heavily on neuroimaging, and sometimes on genetic screening. However, the genetic components in Chinese FTD patients remain largely unknown. Only a few FTD cases with established mutations have been reported in China. This study reported the detailed clinical and neuroimaging features in a Chinese behavioral variant FTD family. The role of MAPT gene mutation in Chinese dementia patients was also reviewed. METHODS: By detailed inquiry of all affected individuals in the family, this study summarized the main clinical features of the disease. Four candidate genes (MAPT, PSEN1, PSEN2, and APP) were screened by direct sequencing. Structural magnetic resonance imaging (MRI), functional imaging of cerebral blood flow with arterial spin-labeled MRI (ASL-MRI), and cerebral metabolism with fluorodeoxyglucose positron emission tomography (FDG-PET) were collected in the proband and healthy mutation carriers. RESULTS: By direct sequencing of candidate genes (MAPT, PSEN1, PSEN2, and APP), this study identified the P301L mutation in the MAPT gene in the proband and three unaffected family members. The phenotype of the affected cases was consistent within the pedigree. In this genetically proven behavioral variant FTD (bvFTD) patient, the maps of hypoperfusion on ASL-MRI look fairly similar to the hypometabolism on FDG-PET. The clinical feature for this bvFTD was in line with the hypoperfusion or hypometabolism pattern on functional neuroimagings. The phenotype of P301L in east Asia seems similar to western countries. CONCLUSION: For the inherited FTD patients, ASL-MRI and genetic identification were strongly recommended for the final diagnosis. In case of being underestimated, the role of MAPT gene mutation in Chinese FTD patients warrants further investigation.

Early Combined Therapy with Pharmacologically Induced Hypothermia and Edaravone Exerts Neuroprotective Effects in a Rat Model of Intracerebral Hemorrhage.

In present study, we evaluated acute neuroprotective effects of combined therapy with pharmacologically induced hypothermia and edaravone in a rat model of intracerebral hemorrhage (ICH). ICH was caused by injection of 0.5 U of collagenase VII to the caudate nucleus of male Sprague-Dawley rats. Sham-treated animals receive injections of normal saline instead of collagenase VII. All animals were randomly divided into five groups: sham group, ICH group, hypothermia group, edavarone (10 mg/kg) group, and combined hypothermia + edavarone group. Hypothermia was induced by injection of the second-generation neurotensin receptor agonist HPI-201 (2 mg/kg at 1 h after ICH; 1 mg/kg at 4 and 7 h after ICH). Hypothermia was sustained for at least 6 h. The study outcomes were the extent of brain edema, permeability of the blood-brain barrier (Evan's blue dye), expression of matrix metalloproteinase-9 and inflammatory cytokines (IL-1ß, IL-4, IL-6, and TNF-α), and expression of apoptosis-related proteins (caspase-3, cytochrome C, Bcl-2, and Bax). Brain edema, permeability of the blood-brain barrier, and expression of metalloproteinase-9 were increased, while expression of caspase-3 and Bcl-2 was decreased by ICH. We observed that the combined therapy was significantly more potent in reverting the above negative trends induced by ICH. In conclusion, our results indicate that a combination of pharmacologically induced hypothermia and edavarone leads to potentiation of their respective neuroprotective effects.

Effects of resveratrol on apoptosis in a rat model of vascular dementia.

Resveratrol is a natural polyphenol widely present in plants, particularly in the skin of red grapes and in wine. It possesses a wide range of biological effects and exhibits neuroprotective effects in numerous diseases. However, data evaluating the effects of resveratrol in vascular dementia (VaD) are lacking. In the present study, the permanent, bilateral common carotid artery occlusion rat model was used to study the effects of resveratrol on VaD. The Morris water maze was used to test the spatial learning and memory performance of the rats. The expression levels of Bax, Bcl-2, cleaved caspase-3 and cleaved poly(ADP-ribose) polymerase (PARP) in the hippocampus were measured. The results showed that resveratrol inhibited memory impairment in the VaD rat model, and attenuated the increases in the expression levels of Bax, cleaved caspase-3 and cleaved PARP and the reductions in the expression levels of Bcl-2 that were induced by VaD. These results provide a novel insight into the neuroprotective effects of resveratrol and its possible therapeutic role in VaD.

Traditional Chinese medicine: a promising candidate for the treatment of Alzheimer's disease.

Alzheimer's disease (AD) is an age-related neurodegenerative disorder, characterized clinically by insidious onset of memory and cognition impairment, emergence of psychiatric symptoms and behavioral disorder, and impairment of activities of daily living (ADL). Traditional Chinese medicine (TCM) is practiced in the Chinese health care system for more than 2,000 years. In recent years, scientists have isolated many novel compounds from herbs, some of which improve dementia with fewer side effects than conventional drugs and are regarded as potential anti-AD drugs. In this review, we summarize the latest research progress on TCM showing their possible role of treatment of AD and other demented diseases and possible pharmacological actions.

Resveratrol improves cognition and reduces oxidative stress in rats with vascular dementia.

Resveratrol possesses beneficial biological effects, which include anti-oxidant, anti-inflammatory and anti-carcinogenic properties. Recently, resveratrol has been shown to exhibit neuroprotective effects in models of Parkinson's disease, cerebral ischemia and Alzheimer's disease. However, its effects on vascular dementia remain unclear. The present study established a rat model of vascular dementia using permanent bilateral common carotid artery occlusion. At 8-12 weeks after model induction, rats were intragastrically administered 25 mg/kg resveratrol daily. Our results found that resveratrol shortened the escape latency and escape distances in the Morris water maze, and prolonged the time spent percentage and swimming distance percentage in the target quadrant during the probe test, indicating that resveratrol improved learning and memory ability in vascular dementia rats. Further experiments found that resveratrol decreased malonyldialdehyde levels, and increased superoxide dismutase activity and glutathione levels in the hippocampus and cerebral cortex of vascular dementia rats. These results confirmed that the neuroprotective effects of resveratrol on vascular dementia were associated with its anti-oxidant properties.

Brain-derived neurotrophic factor prevents beta- amyloid-induced apoptosis of pheochromocytoma cells by regulating Bax/Bcl-2 expression.

Brain-derived neurotrophic factor was utilized in the present study to treat cell injury models induced by aggregated ß-amyloid(25-35). Methylthiazolyldiphenyl-tetrazolium bromide assay and western blot analysis showed that brain-derived neurotrophic factor provided neuroprotection against cellular apoptosis by suppressing the decline in ß-amyloid(25-35)-induced cell activity and the increasing ratio of Bax/Bcl-2. After treating pheochromocytoma cells with tyrosine kinase receptor B receptor inhibitor K252a, brain-derived neurotrophic factor reverses the above-mentioned changes. The experimental findings suggested that brain-derived neurotrophic factor prevented ß-amyloid peptide-induced cellular apoptosis by modulating Bax/Bcl-2 expression, and this effect was associated with binding to the specific tyrosine kinase receptor B receptor.

Involvement of protein trafficking in deprenyl-induced alpha-secretase activity regulation in PC12 cells.

Deprenyl is a selective B-type monoamine oxidase inhibitor and a neuroprotective agent that has been used to slow the progress of Alzheimer's disease for many years. We previously demonstrated that deprenyl could stem amyloid precursor protein processing (APP) toward the non-amyloidogenic pathway through mitogen activated protein kinase (MAPK) and protein kinase C (PKC)-dependent signaling pathways [Yang, H.Q., Ba, M.W., Ren, R.J., Zhang, Y.H., Ma, J.F., Pan, J., Lu, G.Q., Chen, S.D., 2007a. Mitogen activated protein kinase and protein kinase C mediated promotion of sAPPalpha by deprenyl. Neurochem. Int. 50, 74-82.]. The experiment here further showed that deprenyl could increase alpha-secretase activity in a dose-dependent manner in PC12 cells. Deprenyl increased alpha-secretase activity can be partially blocked by pretreatment with brefeldin A, an intracellular protein transport inhibitor, suggesting involvement of protein trafficking in deprenyl regulated alpha-secretase activity. In accordance with this, the experiment showed that brefeldin A also decreased sAPPalpha release induced by deprenyl. Deprenyl promoted ADAM10 transported to the membrane fraction, and this effect was blocked by pretreatment with brefeldin A. The immunocytochemistry staining revealed that deprenyl promoted colocalization of ADAM10 with PKCalpha and PKCepsilon isoforms. These data suggest a novel pharmacological mechanism in which deprenyl increased alpha-secretase activity via protein trafficking related mechanism.

PMS777, a new cholinesterase inhibitor with anti-platelet activated factor activity, regulates amyloid precursor protein processing in vitro.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized clinically by progressive impairment of memory and cognition. Previous data have shown that beta-amyloid (Abeta) cascade plays a central role in AD pathophysiology and thus drugs regulate amyloid precursor protein (APP) metabolism may have therapeutic potential. Here the effects of PMS777, a new cholinesterase inhibitor with anti-platelet activated factor activity, on APP processing were investigated. Using SH-SY5Y(APP695) cells, it showed that PMS777 treatment caused significant decreased secretion of sAPPalpha into the conditioned media without affecting cellular holoAPP synthesis. When PC12 cells were incubated with PMS777, the same effect was observed. The data also indicated that 10 muM PMS777 incubation decreased the release of Abeta42 into the cell media as compared with vehicle group in SH-SY5Y(APP695) cells. Pretreatment of cells with M-receptor scopolamine antagonized the decreased secretion of sAPPalpha induced by PMS777, but N-receptor alpha-bungarotoxin pretreatment did not have such an effect. These results indicated that PMS777 could modulate APP processing in vitro and that decreasing Abeta generation might demonstrate its therapeutic potential in AD.

[Effect of estrogen-depletion and 17beta-estradiol replacement therapy upon rat hippocampus beta-amyloid generation].

OBJECTIVE: To observe the effects of estrogen depletion and 17beta-estradiol replacement therapy upon ratbeta-amyloid (Abeta) generation and the possible related mechanisms. METHODS: Rat ovaries were ectomized to mimic estrogen-depletion models and then 17beta-estradiol was administered by powdering hormone into soy-free chow as a way of replacement therapy. ELISA was carried out to detect rat hippocampus Abeta levels and alpha- and beta-secretase activities were measured after the experiment. The effects of estrogen depletion and 17beta-estradiol replacement therapy upon beta-secretase (BACE1) and neprilysin (NEP) expression were also analyzed by Western blot. RESULTS: Ovariectomy significantly decreased estrogen level [(11 + or - 4) pg/ml, P < 0.01] as compared with control group [(21 + or - 8) pg/ml] while 17beta-estradiol administration increased the estrogen level [(63 + or - 13) pg/ml, P < 0.01] in blood. The Abeta40 [(28.5 + or - 4.5) ng/ml, P < 0.01] and Abeta42 [(4.5 + or - 1.2) ng/ml, P < 0.01] levels were higher in ovariectomy group as compared with their respective control group [with Abeta40 (14.4 + or - 2.4) ng/ml and Abeta42 (2.8 + or - 0.4) ng/ml respectively]. But the effects of ovariectomy on Abeta content can be partially reversed by 17beta-estradiol replacement therapy [with Abeta40 (20.3 + or - 3.2)ng/ml, P < 0.01 and Abeta42 (3.8 + or - 0.5)ng/ml, P < 0.01 respectively]. Estrogen depletion decreased alpha-secretase activity (67.5%, P < 0.01) and increased beta-secretase activity (145.8%, P < 0.01) and this effect can be blocked by 17beta-estradiol administration [with alpha-secretase activity to 90.2% (P < 0.01) and beta-secretase activity to 92.4% (P < 0.01)]. Ovariectomy increased BACE1 expression (135.4%, P < 0.01) and decreased NEP expression (40.8%, P < 0.01) and this effect can be partially antagonized by 17beta-estradiol supplementary [with BACE to 103.5% (P < 0.01) and NEP to 88.4% (P < 0.01)]. CONCLUSION: Estrogen depletion can increase Abeta generation through the effects of increased beta-secretase activity and decreased alpha-secretase activity. Ovariectomy also increases BACE1 expression and decreases NEP expression. The 17beta-estradiol supplementary can decrease Abeta generation and this may to some extent explain why estrogen replacement therapy can decrease the risk of Alzheimer's disease in postmenopausal women.

Economic impact of dementia in developing countries: an evaluation of Alzheimer-type dementia in Shanghai, China.

The main objective of this study was to assess the economic cost of Alzheimer's disease (AD) in Shanghai, China, as a pilot study for future evaluations. Sixty-seven patients with AD were interviewed, and the information of the AD-related cost and resources used was collected from October 2005 to September 2006. By retrospective analysis, annual costs were calculated and expressed in Chinese renminbi (RMB). Direct cost per patient per year averaged approximately 8,432 RMB (1,058 USD), indirect cost per patient per year was 10,568 RMB (1,326 USD), and annual costs were 19,001 RMB (2,384 USD) per patient per year in this investigation. Total cost was significantly associated with the degree of severity including cognitive function (MMSE) and activity of daily living (ADL). With the increase in the number of persons at risk for developing AD, the economic burden of AD patients in China is significantly heavy.

Protective effects of erythropoietin on tau phosphorylation induced by beta-amyloid.

Neuropathological studies have demonstrated that the presence of neurofibrillary tangles (NFTs) is one of the most prominent pathologic characteristics of Alzheimer's disease (AD). The microtubule-associated protein tau is the major component of NFTs, and its abnormal hyperphosphorylation leads to the destabilization of microtubules, impaired axonal transport, and eventual death of the neurons. The hematopoietic cytokine erythropoietin (Epo) is now considered as a viable agent with regard to central nervous system injury in a variety of cellular systems. Here we report that Epo prevented tau hyperphosphorylation in SH-SY5Y cells exposed to the beta-amyloid peptide and that this effect may depend on the PI3K/Akt-GSK-3beta pathway. This study provides new molecular insight into the neuroprotective effect of Epo and suggests its possible therapeutic role in the management of AD.

Expression of FasL and its interaction with Fas are mediated by c-Jun N-terminal kinase (JNK) pathway in 6-OHDA-induced rat model of Parkinson disease.

Our previous studies and those of others have strongly suggested that c-Jun N-terminal kinase (JNK) signaling pathway plays a critical role in 6-hydroxydopamine (6-OHDA)-induced dopaminergic neuron injury in the substantia nigra. However, the downstream mechanism that accounts for the proapoptotic actions of JNK in 6-OHDA lesion remains to be investigated in detail. Fas, a member of the tumor necrosis factor receptor family with proapoptotic functions, was reported to be elevated within the striatum and substantia nigra pars compacta (SNc) of Parkinson's disease (PD) patients. In the present study, we examined the changes in the protein level of Fas ligand (FasL) and its interaction with Fas in a rat model of PD. We demonstrate that the expression of FasL and not Fas was increased after 6-OHDA lesion; additionally, the interaction of FasL and Fas was increased due to 6-OHDA lesion. This indicates that the 6-OHDA-induced activation of Fas signaling pathway is mediated by JNK and that FasL may be a promising target in the therapeutic approach for PD patients.

New protein kinase C activator regulates amyloid precursor protein processing in vitro by increasing alpha-secretase activity.

The beta amyloid (Abeta) cascade has been at the forefront of the hypothesis used to describe the pathogenesis of Alzheimer's disease (AD). It is generally accepted that drugs that can regulate the processing of the amyloid precursor protein (APP) toward the non-amyloidogenic pathway may have a therapeutic potential. Previous studies have shown that protein kinase C (PKC) hypofunction has an important role in AD pathophysiology. Therefore, the effects of a new PKC activator, alpha-APP modulator [(2S,5S)-(E,E)-8-(5-(4-(trifluoromethyl)phenyl)-2,4-pentadienoylamino)benzolactam (TPPB)], on APP processing were investigated. Using PC12 cells and SH-SY5Y(APP695) cells, it was found that TPPB promoted the secretion of sAPPalpha without affecting full-length expression of APP. The increase in sAPPalpha by TPPB was blocked by inhibitors of PKC, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and tyrosine kinase, suggesting the involvement of these signal transduction pathways. TPPB increased alpha-secretase activity [a disintegrin and metalloproteinase (ADAM)10 and 17], as shown by direct fluorescence activity detection and Western blot analysis. TPPB-induced sAPPalpha release was blocked by the metalloproteinase inhibitor TAPI-2, furin inhibitor CMK and by the protein-trafficking inhibitor brefeldin. The results also showed that TPPB decreased beta-secretase activity, Abeta40 release and beta site APP-cleaving enzyme 1 (BACE1) expression, but did not significantly affect neprilysin (NEP) and insulin-degrading enzyme (IDE) expression. Our data indicate that TPPB could direct APP processing towards the non-amyloidogenic pathway by increasing alpha-secretase activity, and suggest its therapeutic potential in AD.