Storage time affects the level and diagnostic efficacy of plasma biomarkers for neurodegenerative diseases

Several promising plasma biomarker proteins,such as amyloid-β(Aβ),tau,neurofilament light chain,and glial fibrillary acidic protein,are widely used for the diagnosis of neurodegenerative diseases.However,little is known about the long-term stability of these biomarker proteins in plasma samples stored at-80°C.We aimed to explore how storage time would affect the diagnostic accuracy of these biomarkers using a large cohort.Plasma samples from 229 cognitively unimpaired individuals,encompassing healthy controls and those experiencing subjective cognitive decline,as well as 99 patients with cognitive impairment,comprising those with mild cognitive impairment and dementia,were acquired from the Sino Longitudinal Study on Cognitive Decline project.These samples were stored at-80°C for up to 6 years before being used in this study.Our results showed that plasma levels of Aβ42,Aβ40,neurofilament light chain,and glial fibrillary acidic protein were not significantly correlated with sample storage time.However,the level of total tau showed a negative correlation with sample storage time.Notably,in individuals without cognitive impairment,plasma levels of total protein and tau phosphorylated protein threonine 181(p-tau181)also showed a negative correlation with sample storage time.This was not observed in individuals with cognitive impairment.Consequently,we speculate that the diagnostic accuracy of plasma p-tau181 and the p-tau181 to total tau ratio may be influenced by sample storage time.Therefore,caution is advised when using these plasma biomarkers for the identification of neurodegenerative diseases,such as Alzheimer's disease.Furthermore,in cohort studies,it is important to consider the impact of storage time on the overall results.

MicroRNAs as potential biomarkers for diagnosis of post-traumatic stress disorder

Post-traumatic stress disorder is a mental disorder caused by exposure to severe traumatic life events.Currently,there are no validated biomarkers or laboratory tests that can distinguish between trauma survivors with and without post-traumatic stress disorder.In addition,the heterogeneity of clinical presentations of post-traumatic stress disorder and the overlap of symptoms with other conditions can lead to misdiagnosis and inappropriate treatment.Evidence suggests that this condition is a multisystem disorder that affects many biological systems,raising the possibility that peripheral markers of disease may be used to diagnose post-traumatic stress disorder.We performed a PubMed search for microRNAs(miRNAs)in post-traumatic stress disorder(PTSD)that could serve as diagnostic biomarkers and found 18 original research articles on studies performed with human patients and published January 2012 to December 2023.These included four studies with whole blood,seven with peripheral blood mononuclear cells,four with plasma extracellular vesicles/exosomes,and one with serum exosomes.One of these studies had also used whole plasma.Two studies were excluded as they did not involve microRNA biomarkers.Most of the studies had collected samples from adult male Veterans who had returned from deployment and been exposed to combat,and only two were from recently traumatized adult subjects.In measuring miRNA expression levels,many of the studies had used microarray miRNA analysis,miRNA Seq analysis,or NanoString panels.Only six studies had used real time polymerase chain reaction assay to determine/validate miRNA expression in PTSD subjects compared to controls.The miRNAs that were found/validated in these studies may be considered as potential candidate biomarkers for PTSD and include miR-3130-5p in whole blood;miR-193a-5p,-7113-5p,-125a,-181c,and-671-5p in peripheral blood mononuclear cells;miR-10b-5p,-203a-3p,-4488,-502-3p,-874-3p,-5100,and-7641 in plasma extracellular vesicles/exosomes;and miR-18a-3p and-7-1-5p in blood plasma.Several important limitations identified in the studies need to be taken into account in future studies.Further studies are warranted with war veterans and recently traumatized children,adolescents,and adults having PTSD and use of animal models subjected to various stressors and the effects of suppressing or overexpressing specific microRNAs.

MicroRNAs as potential diagnostic biomarkers for bipolar disorder

Abnormal expression of microRNAs is connected to brain development and disease and could provide novel biomarkers for the diagnosis and prognosis of bipolar disorder. We performed a PubMed search for microRNA biomarkers in bipolar disorder and found 18 original research articles on studies performed with human patients and published from January 2011 to June 2023. These studies included microRNA profiling in bloodand brain-based materials. From the studies that had validated the preliminary findings,potential candidate biomarkers for bipolar disorder in adults could be miR-140-3p,-30d-5p,-330-5p,-378a-5p,-21-3p,-330-3p,-345-5p in whole blood, miR-19b-3p,-1180-3p,-125a-5p, let-7e-5p in blood plasma, and miR-7-5p,-23b-5p,-142-3p,-221-5p,-370-3p in the blood serum. Two of the studies had investigated the changes in microRNA expression of patients with bipolar disorder receiving treatment. One showed a significant increase in plasma miR-134 compared to baseline after 4 weeks of treatment which included typical antipsychotics, atypical antipsychotics, and benzodiazepines. The other study had assessed the effects of prescribed medications which included neurotransmitter receptorsite binders(drug class B) and sedatives, hypnotics, anticonvulsants, and analgesics(drug class C) on microRNA results. The combined effects of the two drug classes increased the significance of the results for miR-219 and-29c with miR-30e-3p and-526b* acquiring significance. MicroRNAs were tested to see if they could serve as biomarkers of bipolar disorder at different clinical states of mania, depression, and euthymia. One study showed that upregulation in whole blood of miR-9-5p,-29a-3p,-106a-5p,-106b-5p,-107,-125a-3p,-125b-5p and of miR-107,-125a-3p occurred in manic and euthymic patients compared to controls, respectively, and that upregulation of miR-106a-5p,-107 was found for manic compared to euthymic patients. In two other studies using blood plasma,downregulation of miR-134 was observed in manic patients compared to controls, and dysregulation of miR-134,-152,-607,-633,-652,-155 occurred in euthymic patients compared to controls. Finally, microRNAs such as miR-34a,-34b,-34c,-137, and-140-3p,-21-3p,-30d-5p,-330-5p,-378a-5p,-134,-19b-3p were shown to have diagnostic potential in distinguishing bipolar disorder patients from schizophrenia or major depressive disorder patients, respectively. Further studies are warranted with adolescents and young adults having bipolar disorder and consideration should be given to using animal models of the disorder to investigate the effects of suppressing or overexpressing specific microRNAs.

Autism spectrum disorder:difficulties in diagnosis and microRNA biomarkers

We performed a PubMed search for microRNAs in autism spectrum disorder that could serve as diagnostic biomarkers in patients and selected 17 articles published from January 2008 to December 2023,of which 4 studies were performed with whole blood,4 with blood plasma,5 with blood serum,1 with serum neural cell adhesion molecule L1-captured extracellular vesicles,1 with blood cells,and 2 with peripheral blood mononuclear cells.Most of the studies involved children and the study cohorts were largely males.Many of the studies had performed microRNA sequencing or quantitative polymerase chain reaction assays to measure microRNA expression.Only five studies had used real-time polymerase chain reaction assay to validate microRNA expression in autism spectrum disorder subjects compared to controls.The microRNAs that were validated in these studies may be considered as potential candidate biomarkers for autism spectrum disorder and include miR-500a-5p,-197-5p,-424-5p,-664a-3p,-365a-3p,-619-5p,-664a-3p,-3135a,-328-3p,and-500a-5p in blood plasma and miR-151a-3p,-181b-5p,-320a,-328,-433,-489,-572,-663a,-101-3p,-106b-5p,-19b-3p,-195-5p,and-130a-3p in blood serum of children,and miR-15b-5p and-6126 in whole blood of adults.Several important limitations were identified in the studies reviewed,and need to be taken into account in future studies.Further studies are warranted with children and adults having different levels of autism spectrum disorder severity and consideration should be given to using animal models of autism spectrum disorder to investigate the effects of suppressing or overexpressing specific microRNAs as a novel therapy.

MicroRNAs as potential biomarkers for diagnosis of schizophrenia and influence of antipsychotic treatment

Chara cterized by positive symptoms(such as changes in behavior or thoughts,including delusions and hallu cinations),negative symptoms(such as apathy,anhedonia,and social withdrawal),and cognitive impairments,schizophrenia is a chro nic,severe,and disabling mental disorder with late adolescence or early adulthood onset,Antipsychotics are the most commonly used drugs to treat schizophrenia,but those currently in use do not fully reverse all three types of symptoms characte rizing this condition.Schizophrenia is frequently misdiagnosed,resulting in a delay of or inappropriate treatment.Abnormal expression of microRNAs is connected to brain development and disease and could provide novel biomarkers for the diagnosis and prognosis of schizophrenia.The recent studies reviewed included microRNA profiling in blood-and urine-based materials and nervous tissue mate rials.From the studies that had validated the preliminary findings,potential candidate biomarkers for schizophrenia in adults could be miR-22-3p,-30e-5p,-92a-3p,-148b-5p,-181a-3p,-181a-5p,-181b-5p,-199 b-5p,-137 in whole blood,and miR-130b,-193a-3p in blood plasma.Antipsychotic treatment of schizophrenia patients was found to modulate the expression of certain microRNAs including miR-130b,-193a-3p,-132,-195,-30e,-432 in blood plasma.Further studies are warranted with adolescents and young adults having schizophrenia and consideration should be given to using animal models of the disorder to investigate the effect of suppressing or overexpressing specific microRNAs.

Plasma microRNAs as potential new biomarkers for early detection of early gastric cancer

BACKGROUND Early gastric cancer(EGC), compared with advanced gastric cancer(AGC), has a higher 5-year survival rate. However, due to the lack of typical symptoms and the difficulty in diagnosing EGC, no effective biomarkers exist for the detection of EGC, and gastroscopy is the only detection method.AIM To provide new biomarkers with high specificity and sensitivity through analyzed the differentially expressed microRNAs(miRNAs) in EGC and AGC and compared them with those in benign gastritis(BG).METHODSWe examined the differentially expressed miRNAs in the plasma of 30 patients with EGC, AGC, and BG by miRNA chip analysis. Then, we analyzed and selected the significantly different miRNAs using bioinformatics. Reverse transcription quantitative real-time polymerase chain reaction(RT-qPCR)confirmed the relative transcription level of these miRNAs in another 122 patients, including patients with EGC, AGC, Helicobacter pylori(H. pylori)-negative gastritis(Control-1), and H. pylori-positive atrophic gastritis(Control-2).To establish a diagnostic model for the detection of plasma miRNA in EGC, we chose miRNAs that can be used to determine EGC and AGC from Control-1 and Control-2 and miRNAs in EGC from all other groups.RESULTS Among the expression profiles of the miRNA chips in the three groups in the discovery set, of 117 aberrantly expressed miRNAs, 30 confirmed target prediction, whereas 14 were included as potential miRNAs. The RT-qPCR results showed that 14 potential miRNAs expression profiles in the two groups exhibited no differences in terms of H. pylori-negative gastritis(Control-1) and H. pyloripositive atrophic gastritis(Control-2). Hence, these two groups were incorporated into the Control group. A combination of four types of miRNAs,miR-7641, miR-425-5 p, miR-1180-3 p and miR-122-5 p, were used to effectively distinguish the Cancer group(EGC + AGC) from the Control group [area under the curve(AUC) = 0.799, 95% confidence interval(CI): 0.691-0.908, P < 0.001].Additionally, miR-425-5 p, miR-24-3 p, miR-1180-3 p and miR-122-5 p were utilized to distinguish EGC from the Control group(AUC = 0.829, 95%CI: 0.657-1.000, P =0.001). Moreover, the miR-24-3 p expression level in EGC was lower than that in the AGC(AUC = 0.782, 95%CI: 0.571-0.993, P = 0.029), and the miR-4632-5 p expression level in EGC was significantly higher than that in AGC(AUC = 0.791,95%CI: 0.574-1.000, P = 0.024).CONCLUSION The differentially expressed circulatory plasma miR-425-5 p, miR-1180-3 p, miR-122-5 p, miR-24-3 p and miR-4632-5 p can be regarded as a new potential biomarker panel for the diagnosis of EGC. The prediction and early diagnosis of EGC can be considerably facilitated by combining gastroscopy with the use of these miRNA biomarkers, thereby optimizing the strategy for effective detection of EGC. Nevertheless, larger-scale human experiments are still required to confirm our findings.

MicroRNAs as potential biomarkers for diagnosis of attention deficit hyperactivity disorder

Inappropriate levels of hyperactivity,impulsivity,and inattention characterize attention deficit hyperactivity disorder,a common childhood-onset neuropsychiatric disorder.The cognitive function and learning ability of children with attention deficit hyperactivity disorder are affected,and these symptoms may persist to adulthood if they are not treated.The diagnosis of attention deficit hyperactivity disorder is only based on symptoms and objective tests for attention deficit hyperactivity disorder are missing.Treatments for attention deficit hyperactivity disorder in children include medications,behavior therapy,counseling,and education services which can relieve many of the symptoms of attention deficit hyperactivity disorder but cannot cure it.There is a need for a molecular biomarker to distinguish attention deficit hyperactivity disorder from healthy subjects and other neurological conditions,which would allow for an earlier and more accurate diagnosis and appropriate treatment to be initiated.Abnormal expression of microRNAs is connected to brain development and disease and could provide novel biomarkers for the diagnosis and prognosis of attention deficit hyperactivity disorder.The recent studies reviewed had performed microRNA profiling in whole blood,white blood cells,blood plasma,and blood serum of children with attention deficit hyperactivity disorder.A large number of microRNAs were dysregulated when compared to healthy controls and with some overlap between individual studies.From the studies that had included a validation set of patients and controls,potential candidate biomarkers for attention deficit hyperactivity disorder in children could be miR-140-3p,let-7g-5p,-30e-5p,-223-3p,-142-5p,-486-5p,-151a-3p,-151a-5p,and-126-5p in total white blood cells,and miR-4516,-6090,-4763-3p,-4281,-4466,-101-3p,-130a-3p,-138-5p,-195-5p,and-106b-5p in blood serum.Further studies are warranted with children and adults with attention deficit hyperactivity disorder,and consideration should be given to utilizing rat models of attention deficit hyperactivity disorder.Animal studies could be used to confirm microRNA findings in human patients and to test the effects of targeting specific microRNAs on disease progression and behavior.

Screening for potential biomarkers of traditional Chinese medicinechest impediment syndromesbased on plasma metabonomics

OBJECTIVE To have a systematic pathomechanism view of three chest impediment.syndromes of Qi Deficiency and Blood Stasis syndrome(QDBS),Qi Stagnation and Blood Stasis syn.drome(QSBS),Cold Obstruction and Qi Stagnation syndrome(COQS) and further investigate the changed metabolome and related pathways for screening potential biomarkers in rat plasma.METHODS According to clinical pathogeny,three kinds of syndrome models were established to simulate the disease of chest impediment.Plasma metabonomics based on UPLC-Q-TOF/MS was applied in this research to detected small molecule metabolites for identifyingthe special potential biomarkers of three chest impediment syndromes,respectively.RESULTS Significant metabolic differences were observed between thecontrol group and three syndrome groups.Furthermore,three syndrome groups were distinguished clearly by pattern recognition method.The particular metabolites contributing most to the classification of three chest impediment syndromes were identified.In the QSBS group,the potential biomarkers could include 2-keto-glutaramic acid,L-methionine,L-homocysteic acid,octadecanamide,stearoylglycine,behenic acid,linoleylcarnitine,lysoPC(14:1(9 Z)),indoxyl sulfate and cholic acid.In the COQS group,they could be aminoadipic acid,palmitic amide,oleamide,lysoPC(P-16:0),lysoPC(P-18:0),lysoPC(20:2(11 Z,14 Z)),9-HETE and tauroursodeoxycholic acid.Moreover,4-pyridoxic acid,L-palmi.toylcarnitine,lysoPC(20:0),lysoPC(22:5(4 Z,7 Z,10 Z,13 Z,16 Z)),3-hydroxyhexadecanoic acid and arachidonic acid could be the potential biomarkers for the QDBS group.CONCLUSION Three chest impediment syndromes have their own potential biomarkers.Each special metabolite has its owndifferent metabolic pathway.Both metabolismof cysteine and methionine,and metabolism of alanine,aspartate and glutamate are the main pathways in regulation of metabolic disorders in QSBS syndrome.Lysine biosynthesis and degradation,fatty acid metabolism,and glycerophospholipid metabolism are the main pathways in regulation of metabolic disorders in COQS syndrome.Arachidonic acid metabolism,fatty acid metabolism,fatty acid elongation in mitochondria,and vitamin B6 metabolism are the main pathways in regulation of metabolic disorders in QDBS syndrome.These endogenous substances were indicated as the special potential biomarkers for three chest impediment syndromes and worth studying in depth.

MicroRNAs as non-invasive diagnostic biomarkers for gastric cancer:Current insights and future perspectives

Non-invasive diagnostic biomarkers may contribute to an early identification of gastric cancer(GC) and improve the clinical management.Unfortunately,no sensitive and specific screening biomarkers are available yet and the currently available approaches are limited by the nature of the disease.GC is a heterogenic disease with various distinct genetic and epigenetic events that occur during the multifactorial cascade of carcinogenesis.Micro RNAs(mi RNAs) are commonly deregulated in gastric mucosa during the Helicobacter pylori infection and in stepwise manner from chronic gastritis,through preneoplastic conditions such as atrophic gastritis and intestinal metaplasia,to early dysplasia and invasive cancer.Identification of mi RNAs in blood in 2008 led to a great interest on mi RNA-based diagnostic,prognostic biomarkers in GC.In this review,we provide the most recent systematic review on the existing studies related to mi RNAs as diagnostic biomarkers for GC.Here,we systematically evaluate 75 studies related to differential expression of circulating mi RNAs in GC patients and provide novel view on various heterogenic aspects of the existing data and summarize the methodological differences.Finally,we highlight several important aspects crucial to improve the future translational and clinical research in the field.

MicroRNAs as biomarkers of diabetic retinopathy and disease progression

Diabetes mellitus, together with its complications, has been increasing in prevalence worldwide. Its complications include cardiovascular disease(e.g., myocardial infarction, stroke), neuropathy, nephropathy, and eye complications(e.g., glaucoma, cataracts, retinopathy, and macular edema). In patients with either type 1 or type 2 diabetes mellitus, diabetic retinopathy is the leading cause of visual impairment or blindness. It is characterized by progressive changes in the retinal microvasculature. The progression from nonproliferative diabetic retinopathy to a more advanced stage of moderate to severe nonproliferative diabetic retinopathy and proliferative diabetic retinopathy occurs very quickly after diagnosis of mild nonproliferative diabetic retinopathy. The etiology of diabetic retinopathy is unclear, and present treatments have limited effectiveness. Currently diabetic retinopathy can only be diagnosed by a trained specialist, which reduces the population that can be examined. A screening biomarker of diabetic retinopathy with high sensitivity and specificity would aid considerably in identifying those individuals in need of clinical assessment and treatment. The majority of the studies reviewed identified specific microRNAs in blood serum/plasma able to distinguish diabetic patients with retinopathy from those without retinopathy and for the progresion of the disease from nonproliferative diabetic retinopathy to proliferative diabetic retinopathy. In addition,certain microRNAs in vitreous humor were dysregulated in proliferative diabetic retinopathy compared to controls. A very high percentage of patients with diabetic retinopathy develop Alzheimer’s disease. Thus, identifying diabetic retinopathy by measurement of suitable biomarkers would also enable better screening and treatment of those individuals at risk of Alzheimer’s disease.

Hepatocellular carcinoma: Review of disease and tumor biomarkers

Hepatocellular carcinoma(HCC) is a common malignancy and now the second commonest global cause of cancer death. HCC tumorigenesis is relatively silent and patients experience late symptomatic presentation. As the option for curative treatments is limited to early stage cancers, diagnosis in non-symptomatic individuals is crucial. International guidelines advise regular surveillance of high-risk populations but the current tools lack sufficient sensitivity for early stage tumors on the background of a cirrhotic nodular liver. A number of novel biomarkers have now been suggested in the literature, which may reinforce the current surveillance methods. In addition, recent metabonomic and proteomic discoveries have established specific metabolite expressions in HCC, according to Warburg's phenomenon of altered energy metabolism. With clinical validation, a simple and non-invasive test from the serum or urine may be performed to diagnose HCC, particularly benefiting low resource regions where the burden of HCC is highest.

Modulatory effect of pineapple peel extract on lipid peroxidation,catalase activity and hepatic biomarker levels in blood plasma of alcoholinduced oxidative stressed rats

Objective:To investigate the ability of the methanolic extract of pineapple peel to modulate alcohol-induced lipid peroxidation,changes in catalase activities and hepatic biochemical marker levels in blood plasma.Methods:Oxidative stress was induced by oral administration of ethanol(20%w/v) at a dosage of 5 niL/kg bw in rats.After 28 days of treatment,the rats were fasted overnight and sacrificed by cervical dislocation.Blood was collected with a 2 mL syringe by cardiac puncture and was centrifuged at 3000 rpm for 10 min.The plasma was analyzed to evaluate malondialdehyde(MDA),catalase activity,aspartate aminotransferase(AST),alkaline phosphatase(ALP) and alanine aminotransferase(ALT) concentrations.Results:Administration of alcohol caused a drastic increase(87.74%) in MDA level compared with the control.Pineapple peel extract significantly reduced the MDA level by 60.16%at 2.S mL/kg bw.Rats fed alcohol only had the highest catalase activity,treatment with pineapple peel extract at 2.5 mL/kg bw however, reduced the activity.Increased AST,ALP and ALT activities were observed in rats fed alcohol only respectively,treatment with pineapple peel extract drastically reduced their activities. Conclusions:The positive modulation of lipid peroxidation,catalase activities as well as hepatic biomarker levels of blood plasma by the methanolic extract of pineapple peels under alcoholinduced oxidative stress is an indication of its protective ability in the management of alcoholinduced toxicity.

MicroRNAs as disease progression biomarkers and therapeutic targets in experimental autoimmune encephalomyelitis model of multiple sclerosis

Multiple sclerosis is an autoimmune neurodegenerative disease of the central nervous system characterized by pronounced inflammatory infiltrates entering the brain,spinal cord and optic nerve leading to demyelination.Focal demyelination is associated with relapsing-remitting multiple sclerosis,while progressive forms of the disease show axonal degeneration and neuronal loss.The tests currently used in the clinical diagnosis and management of multiple sclerosis have limitations due to specificity and sensitivity.MicroRNAs(miRNAs)are dysregulated in many diseases and disorders including demyelinating and neuroinflammatory diseases.A review of recent studies with the experimental autoimmune encephalomyelitis animal model(mostly female mice 6–12 weeks of age)has confirmed miRNAs as biomarkers of experimental autoimmune encephalomyelitis disease and importantly at the pre-onset(asymptomatic)stage when assessed in blood plasma and urine exosomes,and spinal cord tissue.The expression of certain miRNAs was also dysregulated at the onset and peak of disease in blood plasma and urine exosomes,brain and spinal cord tissue,and at the post-peak(chronic)stage of experimental autoimmune encephalomyelitis disease in spinal cord tissue.Therapies using miRNA mimics or inhibitors were found to delay the induction and alleviate the severity of experimental autoimmune encephalomyelitis disease.Interestingly,experimental autoimmune encephalomyelitis disease severity was reduced by overexpression of miR-146a,miR-23b,miR-497,miR-26a,and miR-20b,or by suppression of miR-182,miR-181c,miR-223,miR-155,and miR-873.Further studies are warranted on determining more fully miRNA profiles in blood plasma and urine exosomes of experimental autoimmune encephalomyelitis animals since they could serve as biomarkers of asymptomatic multiple sclerosis and disease course.Additionally,studies should be performed with male mice of a similar age,and with aged male and female mice.

MicroRNAs as potential biomarkers in temporal lobe epilepsy and mesial temporal lobe epilepsy

Temporal lobe epilepsy is the most common form of focal epilepsy in adults,accounting for one third of all diagnosed epileptic patients,with seizures originating from or involving mesial temporal structures such as the hippocampus,and many of these patients being refractory to treatment with anti-epileptic drugs.Temporal lobe epilepsy is the most common childhood neurological disorder and,compared with adults,the symptoms are greatly affected by age and brain development.Diagnosis of temporal lobe epilepsy relies on clinical examination,patient history,electroencephalographic recordings,and brain imaging.Misdiagnosis or delay in diagnosis is common.A molecular biomarker that could distinguish epilepsy from healthy subjects and other neurological conditions would allow for an earlier and more accurate diagnosis and appropriate treatment to be initiated.Among possible biomarkers of pathological changes as well as potential therapeutic targets in the epileptic brain are micro RNAs.Most of the recent studies had performed micro RNA profiling in body fluids such as blood plasma and blood serum and brain tissues such as temporal cortex tissue and hippocampal tissue.A large number of micro RNAs were dysregulated when compared to healthy controls and with some overlap between individual studies that could serve as potential biomarkers.For example,in adults with temporal lobe epilepsy,possible biomarkers are miR-199a-3p in blood plasma and miR-142-5p in blood plasma and blood serum.In adults with mesial temporal lobe epilepsy,possible biomarkers are miR-153 in blood plasma and miR-145-3p in blood serum.However,in many of the studies involving patients who receive one or several anti-epileptic drugs,the influence of these on micro RNA expression in body fluids and brain tissues is largely unknown.Further studies are warranted with children with temporal lobe epilepsy and consideration should be given to utilizing mouse or rat and non-human primate models of temporal lobe epilepsy.The animal models could be used to confirm micro RNA findings in human patients and to test the effects of targeting specific micro RNAs on disease progression and behavior.

Cerebrospinal fluid and blood biomarkers in Alzheimer’s disease

Due to an ever aging society and growing prevalence of Alzheimer’s disease(AD),the challenge to meet social and health care system needs will become increasingly difficult.Unfortunately,a definite ante mortem diagnosis is not possible.Thus,an early diagnosis and identification of AD patients is critical for promising,early pharmacological interventions as well as addressing health care needs.The most advanced and most reliable markers areβ-amyloid,total tau and phosphorylated tau in cerebrospinal fluid(CSF).In blood,no single biomarker has been identified despite an intense search over the last decade.The most promising approaches consist of a combination of several bloodbased markers increasing the reliability,sensitivity and specificity of the AD diagnosis.However,contradictory data make standardized testing methods in longitudinal and multi-center studies extremely difficult.In this review,we summarize a range of the most promising CSF and blood biomarkers for diagnosing AD.

MicroRNAs as biomarkers in glaucoma and potential therapeutic targets

Glaucoma is a neurodegenerative disease in which optic nerve damage and visual field defects occur.It is a leading cause of irreversible blindness.Its pathogenesis is largely unknown although several risk factors have been identified,with an increase in intraocular pressure being the main one.Lowering of intraocular pressure is the only treatment available.Open-angle glaucoma is the most common form of the condition,accounting for~90%of all cases of glaucoma,with primary open-angle glaucoma and exfoliation glaucoma being the most frequent types.There are strong indications that microRNAs play important roles in the pathogenesis of primary open-angle glaucoma.Most of the recent studies reviewed had performed microRNA profiling in aqueous humor from glaucoma patients compared to controls who were chiefly cataract patients.A very large number of microRNAs were dysregulated but with limited overlap between individual studies.MiRNAs in aqueous humor that could be possible targets for therapeutic intervention are miR-143-3p,miR-125b-5p,and miR-1260b.No ove rlap of findings occurred within the dysregulated miRNAs for blood plasma,blood serum,peripheral blood mononuclear cells,and tears of primary open-angle glaucoma patients.Seve ral impo rtant limitations were identified in these studies.Further studies are warranted of mic roRNA expression in aqueous humor and blood samples of primary open-angle glaucoma patients in the early stages of the disease so that validated biomarkers can be identified and treatment initiated.In addition,whether modifying the levels of specific microRNAs in aqueous humor or tears has a beneficial effect on intraocular pressure and ophthalmic examination of the eyes should be investigated using suitable animal models of glaucoma.

Identification of plasma microRNA-21 as a biomarker for early detection and chemosensitivity of non-small cell lung cancer

Studies have shown cell-free microRNA(miRNA) circulating in the serum and plasma with specific expression in cancer,indicating the potential of using miRNAs as biomarkers for cancer diagnosis and therapy.This study was to investigate whether plasma miRNA-21(miR-21) can be used as a biomarker for the early detection of non-small cell lung cancer(NSCLC) and to explore its association with clinicopathologic features and sensitivity to platinum-based chemotherapy.We used real-time RT-PCR to investigate the expression of miR-21 in the plasma of 63 NSCLC patients and 30 healthy controls and correlated the findings with early diagnosis,pathologic parameters,and treatment.Thirty-five patients(stages IIIB and IV) were evaluable for chemotherapeutic responses:11 had partial response(PR);24 had stable and progressive disease(SD+PD).Plasma miR-21 was significantly higher in NSCLC patients than in age-and sex-matched controls(P<0.001).miR-21 was related to TNM stage(P<0.001),but not related to age,sex,smoking status,histological classification,lymph node status,and metastasis(all P>0.05).This marker yielded a receiver operating characteristic(ROC) curve area of 0.775(95% CI:0.681-0.868) with 76.2% sensitivity and 70.0% specificity.Importantly,miR-21 plasma levels in PR samples were several folds lower than that in SD plus PD samples(P=0.049),and were close to that in healthy controls(P=0.130).Plasma miR-21 can serve as a circulating tumor biomarker for the early diagnosis of NSCLC and is related to the sensitivity to platinum-base chemotherapy.

MicroRNAs as diagnostic and prognostic biomarkers of age-related macular degeneration:advances and limitations

A main cause of vision loss in the elderly is age-related macular degeneration(AMD).Among the cellular,biochemical,and molecular changes linked to this disease,inflammation and angiogenesis appear as being crucial in AMD pathogenesis and progression.There are two forms of the disease:dry AMD,accounting for 80–90%of cases,and wet AMD.The disease usually begins as dry AMD associated with retinal pigment epithelium and photoreceptor degeneration,whereas wet AMD is associated with choroidal neovascularization resulting in severe vision impairment.The new vessels are largely malformed,leading to blood and fluid leakage within the disrupted tissue,which provokes inflammation and scar formation and results in retinal damage and detachment.Micro RNAs are dysregulated in AMD and may facilitate the early detection of the disease and monitoring disease progression.Two recent reviews of micro RNAs in AMD had indicated weaknesses or limitations in four earlier investigations.Studies in the last three years have shown considerable progress in overcoming some of these concerns and identifying specific micro RNAs as biomarkers for AMD.Further large-scale studies are warranted using appropriate statistical methods to take into account gender and age disparity in the study populations and confounding factors such as smoking status.

Biomarkers for hepatocellular cancer

Hepatocellular carcinoma(HCC)is the third leading cause of cancer-related deaths worldwide.If diagnosed early,curative treatment options such as surgical resection,loco-regional therapies,and liver transplantation are available to patients,increasing their chances of survival and improving their quality of life.Unfortunately,most patients are diagnosed with late stage HCC where only palliative treatment is available.Therefore,biomarkers which could detect HCC early with a high degree of sensitivity and specificity,may play a crucial role in the diagnosis and management of the disease.This review will aim to provide an overview of the different biomarkers of HCC comprising those used in the diagnosis of HCC in at risk populations,as well as others with potential for prognosis,risk predisposition and prediction of response to therapeutic intervention.

Serum outperforms plasma in small extracellular vesicle microRNA biomarker studies of adenocarcinoma of the esophagus

BACKGROUND Circulating microRNAs(miRNAs)are potential biomarkers for many diseases.However,they can originate from non-disease specific sources,such as blood cells,and compromise the investigations for miRNA biomarkers.While small extracellular vesicles(sEVs)have been suggested to provide a purer source of circulating miRNAs for biomarkers discovery,the most suitable blood sample for sEV miRNA biomarker studies has not been defined.AIM To compare the mi RNA profiles between matched serum and plasma s EV preparations to determine their suitability for biomarker studies.METHODS Matched serum and plasma samples were obtained from 10 healthy controls and10 patients with esophageal adenocarcinoma.s EV isolates were prepared from serum and plasma using Exo Quick TM and quantified using Nano Sight.RNA was extracted from s EV preparations with the mi RNeasy Serum/Plasma kit and profiled using the Taqman Openarray q PCR.The overall mi RNA content and theexpression of specific mi RNAs of reported vesicular and non-vesicular origins were compared between serum and plasma s EV preparations.The diagnostic performance of a previously identified multi-mi RNA biomarker panel for esophageal adenocarcinoma was also compared.RESULTS The overall mi RNA content was higher in plasma s EV preparations(480 mi RNAs)and contained 97.5%of the mi RNAs found in the serum s EV preparations(412 mi RNAs).The expression of commonly expressed mi RNAs was highly correlated(Spearman’s R=0.87,P<0.0001)between the plasma and serum s EV preparations,but was consistently higher in the plasma s EV preparations.Specific blood-cell mi RNAs(hsa-mi R-223-3 p,hsa-mi R-451 a,mi R-19 b-3 p,hsa-mi R-17-5 p,hsa-mi R-30 b-5 p,hsa-mi R-106 a-5 p,hsa-mi R-150-5 p and hsa-mi R-92 a-3 p)were expressed at 2.7 to 9.6 fold higher levels in the plasma s EV preparations compared to serum s EV preparations(P<0.05).In plasma s EV preparations,the percentage of protein-associated mi RNAs expressed at relatively higher levels(Ct 20-25)was greater than serum s EV preparations(50%vs 31%).While the percentage of vesicle-associated mi RNAs expressed at relatively higher levels was greater in the serum s EV preparations than plasma s EV preparations(70%vs 44%).A 5-mi RNA biomarker panel produced a higher cross validated accuracy for discriminating patients with esophageal adenocarcinoma from healthy controls using serum s EV preparations compared with plasma s EV preparations(AUROC 0.80 vs 0.54,P<0.05).CONCLUSION Although plasma s EV preparations contained more mi RNAs than serum s EV preparations,they also contained more mi RNAs from non-vesicle origins.Serum appears to be more suitable than plasma for s EV mi RNAs biomarkers studies.