C-ter100 peptide derived from Vibrio vEP-45 protease acts as a pathogen-associated molecular pattern to induce inflammation and innate immunity.

The bacterium Vibrio vulnificus causes fatal septicemia in humans. Previously, we reported that an extracellular metalloprotease, vEP-45, secreted by V. vulnificus, undergoes self-proteolysis to generate a 34 kDa protease (vEP-34) by losing its C-terminal domain to produce the C-ter100 peptide. Moreover, we revealed that vEP-45 and vEP-34 proteases induce blood coagulation and activate the kallikrein/kinin system. However, the role of the C-ter100 peptide fragment released from vEP-45 in inducing inflammation is still unclear. Here, we elucidate, for the first time, the effects of C-ter100 on inducing inflammation and activating host innate immunity. Our results showed that C-ter100 could activate NF-κB by binding to the receptor TLR4, thereby promoting the secretion of inflammatory cytokines and molecules, such as TNF-α and nitric oxide (NO). Furthermore, C-ter100 could prime and activate the NLRP3 inflammasome (NLRP3, ASC, and caspase 1), causing IL-1ß secretion. In mice, C-ter100 induced the recruitment of immune cells, such as neutrophils and monocytes, along with histamine release into the plasma. Furthermore, the inflammatory response induced by C-ter100 could be effectively neutralized by an anti-C-ter100 monoclonal antibody (C-ter100Mab). These results demonstrate that C-ter100 can be a pathogen-associated molecular pattern (PAMP) that activates an innate immune response during Vibrio infection and could be a target for the development of antibiotics.

Alzheimer's disease risk associated with changes in Epstein-Barr virus nuclear antigen 1-specific epitope targeting antibody levels.

BACKGROUND: Alzheimer's disease (AD) is a neurodegenerative disorder influenced by age, sex, genetic factors, immune alterations, and infections. Multiple lines of evidence suggest that changes in antibody response are linked to AD pathology. METHODS: To elucidate the mechanisms underlying AD development, we investigated antibodies that target autoimmune epitopes using high-resolution epitope microarrays. Our study compared two groups: individuals with AD (n = 19) and non-demented (ND) controls (n = 19). To validate the results, we measured antibody levels in plasma samples from AD patients (n = 96), mild cognitive impairment (MCI; n = 91), and ND controls (n = 97). To further explore the invlovement of EBV, we performed epitope masking immunofluorescence microscopy analysis and tests to induce lytic replication using the B95-8 cell line. RESULTS: In this study, we analyzed high-resolution epitope-specific serum antibody levels in AD, revealing significant disparities in antibodies targeting multiple epitopes between the AD and control groups. Particularly noteworthy was the significant down-regulation of antibody (anti-DG#29) targeting an epitope of Epstein-Barr virus nuclear antigen 1 (EBNA1). This down-regulation increased AD risk in female patients (odds ratio up to 6.6), but not in male patients. Our investigation further revealed that the down-regulation of the antibody (anti-DG#29) is associated with EBV reactivation in AD, as indicated by the analysis of EBV VCA IgG or IgM levels. Additionally, our data demonstrated that the epitope region on EBNA1 for the antibody is hidden during the EBV lytic reactivation of B95-8 cells. CONCLUSION: Our findings suggest a potential relationship of EBV in the development of AD in female. Moreover, we propose that antibodies targeting the epitope (DG#29) of EBNA1 could serve as valuable indicators of AD risk in female.

Diagnostic Blood Biomarkers in Alzheimer's Disease.

Potential biomarkers for Alzheimer's disease (AD) include amyloid ß1-42 (Aß1-42), t-Tau, p-Tau181, neurofilament light chain (NFL), and neuroimaging biomarkers. Their combined use is useful for diagnosing and monitoring the progress of AD. Therefore, further development of a combination of these biomarkers is essential. We investigated whether plasma NFL/Aß1-42 can serve as a plasma-based primary screening biomarker reflecting brain neurodegeneration and amyloid pathology in AD for monitoring disease progression and early diagnosis. We measured the NFL and Aß1-42 concentrations in the CSF and plasma samples and performed correlation analysis to evaluate the utility of these biomarkers in the early diagnosis and monitoring of AD spectrum disease progression. Pearson's correlation analysis was used to analyse the associations between the fluid biomarkers and neuroimaging data. The study included 136 participants, classified into five groups: 28 cognitively normal individuals, 23 patients with preclinical AD, 22 amyloid-negative patients with amnestic mild cognitive impairment, 32 patients with prodromal AD, and 31 patients with AD dementia. With disease progression, the NFL concentrations increased and Aß1-42 concentrations decreased. The plasma and CSF NFL/Aß1-42 were strongly correlated (r = 0.558). Plasma NFL/Aß1-42 was strongly correlated with hippocampal volume/intracranial volume (r = 0.409). In early AD, plasma NFL/Aß1-42 was associated with higher diagnostic accuracy than the individual biomarkers. Moreover, in preclinical AD, plasma NFL/Aß1-42 changed more rapidly than the CSF t-Tau or p-Tau181 concentrations. Our findings highlight the utility of plasma NFL/Aß1-42 as a non-invasive plasma-based biomarker for early diagnosis and monitoring of AD spectrum disease progression.

Effect of Vibrio-Derived Extracellular Protease vEP-45 on the Blood Complement System.

Vibrio vulnificus is a pathogenic bacterium that can causes wound infections and fetal septicemia. We have reported that V. vulnificus ATCC29307 produces an extracellular zinc-metalloprotease (named vEP-45). Our previous results showed that vEP-45 can convert prothrombin to active thrombin and also activate the plasma kallikrein/kinin system. In this study, the effect of vEP-45 on the activation of the complement system was examined. We found that vEP-45 could proteolytically convert the key complement precursor molecules, including C3, C4, and C5, to their corresponding active forms (e.g., C3a, C3b, C4a, C4b, and C5a) in vitro cleavage assays. C5b production from C5 cleavage mediated by vEP-45 was not observed, whereas the level of C5a was increased in a dose-dependent manner compared to that of the non-treated control. The cleavage of the complement proteins in human plasma by vEP-45 was also confirmed via Western blotting. Furthermore, vEP-45 could convert C3 and C5 to active C3a and C5a as a proinflammatory mediator, while no cleavage of C4 was observed. These results suggest that vEP-45 can activate the complement system involved in innate immunity through an alternative pathway.

Regulation of p53 Activity by (+)-Epiloliolide Isolated from Ulva lactuca.

Ulva lactuca (U. lactuca) is a green alga distributed worldwide and used as a food and cosmetic material. In our previous study, we determined the effects of U. lactuca methanol extracts on the UVB-induced DNA repair. In the present study, we fractionated U. lactuca methanol extracts to identify the effective compound for the DNA repair. MTT assay demonstrated that (+)-epiloliolide showed no cytotoxicity up to 100 µM in BJ-5ta human dermal fibroblast. Upon no treatment, exposure to UVB 400 J/m2 decreased cell viability by 45%, whereas (+)-epiloliolide treatment for 24 h after UVB exposure significantly increased the cell viability. In GO and GESA analysis, a number of differentially expressed genes were uniquely expressed in (+)-epiloliolide treated cells, which were enriched in the p53 signaling pathway and excision repair. Immunofluorescence demonstrated that (+)-epiloliolide increased the nuclear localization of p53. Comet assay demonstrated that (+)-epiloliolide decreased tail moment increased by UVB. Western blot analysis demonstrated that (+)-epiloliolide decreased the levels of p-p53, p21, Bax, and Bim, but increased that of Bcl-2. Reverse transcription PCR (RT-PCR) demonstrated that (+)-epiloliolide decreased the levels of MMP 1, 9, and 13, but increased that of COL1A1. These results suggest that (+)-epiloliolide regulates p53 activity and has protective effects against UVB.

Effect of 7-Methylsulfinylheptyl Isothiocyanate on the Inhibition of Melanogenesis in B16-F1 Cells.

Skin aging, characterized by hyperpigmentation, inflammation, wrinkles, and skin cancer, is influenced by intrinsic and extrinsic factors with synergistic effects. Autophagy maintains the homeostatic balance between the degradation, synthesis, and recycling of cellular proteins and organelles, and plays important roles in several cellular and biological processes, including aging. The compound 7-methylsulfinylheptyl isothiocyanate (7-MSI) is a sulfur-containing phytochemical produced by various plants, particularly cruciferous vegetables, with reported anti-inflammatory properties and a role in pathogen defense; however, its effects on skin whitening have not been studied in detail. The purpose of this study was to observe the effects of 7-MSI on skin whitening and autophagy in cultured murine melanoma (B16-F1) cells. Western blotting was used to evaluate the impact of 7-MSI on melanogenesis-, tyrosinase-, and autophagy-associated proteins. The levels of the melanogenesis-associated protein's microphthalmia-associated transcription factor (MITF) and tyrosinase and tyrosinase-related protein-1 were decreased by treatment with 7-MSI under melanogenesis induction. Melanin synthesis also decreased by approximately 63% after treatment with 7-MSI for 73 h, compared with that non-treated controls. In addition, autophagosome formation and the expression levels of the autophagy-related proteins mTOR, p-mTOR, Beclin-1, Atg12, and LC3 were higher in 7-MSI-treated B16-F1 cells than in non-treated cells. These results indicate that 7-MSI can inhibit melanin synthesis in B16-F1 cells by suppressing melanogenesis and autophagy activation and thus can potentially be used as a novel multifunctional cosmetic agent.

Plasma contact factors as novel biomarkers for diagnosing Alzheimer's disease.

BACKGROUND: Alzheimer's disease (AD) is the most common cause of dementia and most of AD patients suffer from vascular abnormalities and neuroinflammation. There is an urgent need to develop novel blood biomarkers capable of diagnosing Alzheimer's disease (AD) at very early stage. This study was performed to find out new accurate plasma diagnostic biomarkers for AD by investigating a direct relationship between plasma contact system and AD. METHODS: A total 101 of human CSF and plasma samples from normal and AD patients were analyzed. The contact factor activities in plasma were measured with the corresponding specific peptide substrates. RESULTS: The activities of contact factors (FXIIa, FXIa, plasma kallikrein) and FXa clearly increased and statistically correlated as AD progresses. We present here, for the first time, the FXIIa cut-off scores to as: > 26.3 U/ml for prodromal AD [area under the curve (AUC) = 0.783, p < 0.001] and > 27.2 U/ml for AD dementia (AUC = 0.906, p < 0.001). We also describe the cut-off scores from the ratios of CSF Aß1-42 versus the contact factors. Of these, the representative ratio cut-off scores of Aß1-42/FXIIa were to be: < 33.8 for prodromal AD (AUC = 0.965, p < 0.001) and < 27.44 for AD dementia (AUC = 1.0, p < 0.001). CONCLUSION: The activation of plasma contact system is closely associated with clinical stage of AD, and FXIIa activity as well as the cut-off scores of CSF Aß1-42/FXIIa can be used as novel accurate diagnostic AD biomarkers.

Two Opposing Roles of SARS-CoV-2 RBD-Reactive Antibodies in Pre-Pandemic Plasma Samples From Elderly People in ACE2-Mediated Pseudovirus Infection.

A novel coronavirus designated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged and caused an outbreak of unusual viral pneumonia. Several reports have shown that cross-reactive antibodies against SARS-CoV-2 also exist in people unexposed to this virus. However, the neutralizing activity of cross-reactive antibodies is controversial. Here, we subjected plasma samples from SARS-CoV-2-unexposed elderly Korean people (n = 119) to bead-based IgG antibody analysis. SARS-CoV-2 S1 subunit-reactive IgG antibody analysis detected positive signals in some samples (59 of 119, 49.6%). SARS-CoV-2 receptor-binding domain (RBD)-reactive antibody levels were most significantly correlated with human coronavirus-HKU1 S1 subunit-reactive antibody levels. To check the neutralizing activity of plasma samples, the SARS-CoV-2 spike pseudotype neutralizing assay was used. However, the levels of cross-reactive antibodies did not correlate with neutralizing activity. Instead, SARS-CoV-2 pseudovirus infection was neutralized by some RBD-reactive plasma samples (n = 9, neutralization ≥ 25%, P ≤ 0.05), but enhanced by other RBD-reactive plasma samples (n = 4, neutralization ≤ -25%, P ≤ 0.05). Interestingly, the blood plasma groups with enhancing and neutralizing effects had high levels of SARS-CoV-2 RBD-reactive antibodies than the plasma group that had no effect. These results suggest that some SARS-CoV-2 RBD-reactive antibodies from pre-pandemic elderly people exert two opposing functions during SARS-CoV-2 pseudovirus infection. In conclusion, preformed RBD-reactive antibodies may have two opposing functions, namely, protecting against and enhancing viral infection. Analysis of the epitopes of preformed antibodies will be useful to elucidate the underlying mechanism.

Alpha-neoendorphin can reduce UVB-induced skin photoaging by activating cellular autophagy.

Skin aging is influenced by several genetic, physiological, and environmental factors. In particular, ultraviolet (UV) exposure is an important factor involved in inducing skin photoaging. Autophagy controlling homeostatic balance between the synthesis, degradation, and recycling of cellular organelles and proteins plays important regulatory roles in several biological processes, including aging. The opioid neuropeptide α-neoendorphin (named NEP) is an endogenous decapeptide (N-YGGFLRKYPK-C) that activates the kappa opioid receptor and exhibits certain anti-aging and anti-wrinkling effects on skin cells; however, its action mechanism has not yet been elucidated. Therefore, the aim of this study was to determine the effects of NEP on anti-skin aging and autophagy activation in human dermal fibroblast cells. Western blot results showed that NEP down-regulates the production of phospho-mammalian target of rapamycin (p-mTOR), whereas increases the expression of key autophagy-related molecules such as Beclin-1, Atg5-Atg12, and LC3-II. The immunocytochemical analysis performed with anti-LC3-II antibody also showed that the autophagic indicators, autophagosomes are formed by NEP. These results suggest that NEP can activate cellular autophagy through mTOR-Beclin-1-mediated signaling pathway. It was also revealed by CM-H2DCF-DA assay and Western blottings that NEP can reduce the production of ultraviolet B (UVB)-induced reactive oxygen species (ROS) like with N-acetylcysteine (NAC), resulting in decreasing the expression levels of skin aging-related proteins, such as phospho-ERK (p-ERK), phospho-p38 (p-p38), and phospho-JNK (p-JNK). Furthermore, NEP could increase the type I procollagen production, while decreasing MMP-1, MMP-2, and MMP-9 activities. Taken together, the results demonstrate that NEP can reduce UVB-induced photoaging by activating autophagy.

Cloning, purification and characterization of a recombinant protease with novel thrombolytic activity in human plasma and rat thrombosis models.

BACKGROUND: There is a need to identify and develop novel thrombolytic agents that can directly digest fibrin clots from various biological resources. OBJECTIVE: To clone, express, purify, and characterize a recombinant protease named rvFMP capable of cleaving fibrinogen, fibrin polymer, and cross-linked fibrin in human plasma milieu and rat thrombosis model systems. RESULTS: We cloned a vFMP-encoding gene from the genomic DNA of V. furnissii KCCM41679 using polymerase chain reaction (PCR), expressed in Escherichia coli, and purified rvFMP (stands for recombinant vibrio furnissii metalloprotease). The proteolytic activity of purified rvFMP enzyme could be clearly inhibited by 1,10-phenanthroline and ethylene glycol tetraacetic acid, but not by diisopropyl fluorophosphate, suggesting that it can be a typical metalloprotease. rvFMP showed an effective proteolytic activity in cleaving cross-linked fibrins in human plasma milieu. Remarkably, rvFMP exhibited a clear thrombolytic activity in rat thrombosis models such as ferric chloride-exposed rat carotid artery and carrageenan-treated rat tail. However, rvFMP (1.5 mg/kg) evoked no internal bleeding and also showed no lethal effect in mice. The recombinant enzyme also showed no cytotoxicity and had an inability to induce tumour necrosis factor-α (TNF-α) in Raw264.7 cells. CONCLUSION: rvFMP can be a candidate enzyme capable of being developed as a novel direct-acting thrombolytic agent.

Comparison of Two Analytical Platforms in Cerebrospinal Fluid Biomarkers for the Classification of Alzheimer's Disease Spectrum with Amyloid PET Imaging.

BACKGROUND: Cerebrospinal fluid (CSF) amyloid-ß1-42 (Aß1-42), total tau protein (t-Tau), and phosphorylated Tau (p-Tau) are ATN biomarkers for Alzheimer's disease (AD) and reflect pathogenic changes in the brain. CSF biomarkers of AD are considered for inclusion in the diagnostic criteria for research and clinical purposes to reduce the uncertainty of clinical diagnosis and to distinguish among AD stages. OBJECTIVE: This study aims to compare two commercially available analytical platforms with respect to accuracy and the potential for early diagnosis of AD. METHODS: A total of 211 CSF samples from healthy control (HC) and AD subjects were analyzed using two analytical platforms, INNOTEST ELISA and INNOBIA AlzBio3 xMAP kits. The accuracy of diagnosis and AUC values distinguishing study groups were compared between the two analytical platforms. RESULTS: The absolute values for Aß1-42, t-Tau, and p-Tau181 levels differed between the two platforms. The Aß1-42 levels decreased, while t-Tau and p-Tau levels increased according to the AD stages. The AUC of Aß1-42 and t-Tau, which distinguish the early stages of AD (preclinical and prodromal AD), were similar between the two platforms, whereas there were significant differences in p-Tau AUC values. CSF p-Tau using the INNOBIA was highly accurate for distinguishing both preclinical AD (AUC = 0.826, cut-off score≥38.89) and prodromal AD (AUC = 0.862, cut-off score≥41.88) from HC. CONCLUSION: The accuracy of CSF p-Tau levels in the preclinical and prodromal AD is higher for the INNOBIA than the INNOTEST, and the early stage AD can be accurately distinguished from HC.

Effectiveness of training program combining chakrayoga and meditation.

Background This study was designed to examine the effectiveness of program combining chakrayoga and meditation on the physical health and disease-related factors and psychological factors of people. Methods Ninety-seven subjects (32-83 years old) who had free from prior experiences in meditation programs or Chakrayoga training courses were assigned to either the experimental group (EXP) (45 subjects; 13 male subjects and 32 female subjects; average age of 60.67 years, SD=11.09 years) or the control group (CONT) of remaining subjects (52 subjects; 14 male subjects and 38 female subjects; average age of 61.58 years, SD=9.70 years). Subjects in the EXP participated in the Chakrayoga Meditation Program for twice a week for 2 h during 6 weeks in each session consisted of 1 h of Chakrayoga and 1 h of meditation. The measurements in this study included the mindfulness, stress response, subjective quality of life, medical symptom checklist, difficulty in emotional regulation and objective of life and sense of control. Results Results revealed that participants in the EXP reported significantly more relief of mindfulness, stress response, subjective quality of life and medical symptom checklist than those in the CONT. Conclusions These findings provide evidence that the Chakrayoga Meditation Program can help relieve the physical health and disease-related factors and psychological factors.

APOE Promoter Polymorphism-219T/G is an Effect Modifier of the Influence of APOE ε4 on Alzheimer's Disease Risk in a Multiracial Sample.

Variants in the APOE gene region may explain ethnic differences in the association of Alzheimer's disease (AD) with ε4. Ethnic differences in allele frequencies for three APOE region SNPs (single nucleotide polymorphisms) were identified and tested for association in 19,398 East Asians (EastA), including Koreans and Japanese, 15,836 European ancestry (EuroA) individuals, and 4985 African Americans, and with brain imaging measures of cortical atrophy in sub-samples of Koreans and EuroAs. Among ε4/ε4 individuals, AD risk increased substantially in a dose-dependent manner with the number of APOE promoter SNP rs405509 T alleles in EastAs (TT: OR (odds ratio) = 27.02, p = 8.80 × 10-94; GT: OR = 15.87, p = 2.62 × 10-9) and EuroAs (TT: OR = 18.13, p = 2.69 × 10-108; GT: OR = 12.63, p = 3.44 × 10-64), and rs405509-T homozygotes had a younger onset and more severe cortical atrophy than those with G-allele. Functional experiments using APOE promoter fragments demonstrated that TT lowered APOE expression in human brain and serum. The modifying effect of rs405509 genotype explained much of the ethnic variability in the AD/ε4 association, and increasing APOE expression might lower AD risk among ε4 homozygotes.

Cerebrospinal Fluid Biomarkers for the Diagnosis of Prodromal Alzheimer's Disease in Amnestic Mild Cognitive Impairment.

BACKGROUND/AIMS: Disease-modifying therapy for Alzheimer's disease (AD) has led to a need for biomarkers to identify prodromal AD and very early stage of AD dementia. We aimed to identify the cutoff values of cerebrospinal fluid (CSF) biomarkers for detecting prodromal AD. METHODS: We assessed 56 patients with amnestic mild cognitive impairment (aMCI) who underwent lumbar puncture. Additionally, 87 healthy elderly individuals and 34 patients with AD dementia served as controls. Positron emission tomography was performed using florbetaben as a probe. We analyzed the concentration of Aß1-42, total tau protein (t-Tau), and tau protein phosphorylated at threonine 181 (p-Tau181) in CSF with INNOTEST enzyme-linked immunosorbent assay. RESULTS: For the detection of prodromal AD in patients with aMCI, the cutoff values of CSF Aß1-42, t-Tau, and p-Tau181 were 749.5 pg/mL, 225.6 pg/mL, and 43.5 pg/mL, respectively. To discriminate prodromal AD in patients with aMCI, the t-Tau/Aß1-42 and -p-Tau181/Aß1-42 ratios defined cutoff values at 0.298 and 0.059, respectively. CONCLUSIONS: CSF biomarkers are very useful tools for the differential diagnosis of prodromal AD in aMCI patients. The concentration of CSF biomarkers is well correlated with the stages of the AD spectrum.

High-throughput epitope profiling of antibodies in the plasma of Alzheimer's disease patients using random peptide microarrays.

The symptoms of Alzheimer's disease (AD), a major cause of dementia in older adults, are linked directly with neuronal cell death, which is thought to be due to aberrant neuronal inflammation. Autoantibodies formed during neuronal inflammation show excellent stability in blood; therefore, they may be convenient blood-based diagnostic markers of AD. Here, we performed microarray analysis of 29,240 unbiased random peptides to be used for comprehensive screening of AD-specific IgG and IgM antibodies in the blood. The results showed that (1) sequence-specific and isotype-specific antibodies are regulated differentially in AD, and combinations of these antibodies showing high area under the receiver operating characteristic curve values (0.862-0.961) can be used to classify AD, (2) AD-specific IgG antibodies arise from IgM antibody-secreting cells that existed before disease onset and (3) target protein profiling of the antibodies identified some AD-related proteins, some of which are involved in AD-related signalling pathways. Therefore, we propose that these epitopes may facilitate the development of biomarkers for AD diagnosis and form the basis for a mechanistic study related to AD progression.

Upregulation of autophagy by Ginsenoside Rg2 in MCF-7 cells.

Autophagy is a major intracellular degradation process that plays an important role in cell survival, stress responses, nutrient sensing and development. Our previous studies have shown that Rg2, a triterpenoid saponin contained in ginseng, protects cells against UVB-induced genotoxicity by increasing DNA repair, in possible association with modulation of protein levels involved in p53 pathway. In this study, we determined an upregulation of autophagy by Rg2. Rg2 treatment for 24 h in MCF-7, a breast cancer cell, did not show cytotoxicity up to 200 µM. Rg2 also upregulated the level of p-p53, p-AMPK, p-ACC, Atg-7 and LC3-II and decreased the level of p62 in concentration-dependent manners. We also determined the level of p53, AMPK, p62, Atg-7 and LC3 after UVB exposure and subsequent incubation in growth medium for 24 h. UVB increased the level of p-p53, p-AMPK, p-ACC and decreased the levels of p62, Atg-7 and LC3-II. Interestingly, Rg2 treatment for 24 h after UVB exposure increased the levels of p-p53, p-AMPK, p-ACC, Atg-7 and LC3-II and decreased the level of cyclobutane pyrimidine dimer, a UVB-induced DNA damage in concentration-dependent manners. All these results suggest that Rg2 increased autophagy and decreased UVB-induced DNA damage, in possible association with the modulation of protein levels in p53- and autophagic pathways.

Protective effects of ginsenoside Rg2 and astaxanthin mixture against UVB-induced DNA damage.

Ultraviolet B (UVB) radiation induces skin damage, skin matrix degradation, and wrinkle formation through photochemical reaction and oxidative stress. Therefore, protecting the skin from UVB can prevent skin aging. In this study, we investigated the effects of a mixture (RA) of Rg2, a ginsenoside, and astaxanthin, an antioxidant, on the responses of HaCaT cells exposed to UVB (700 J/m2). The cells were incubated for 24 h after UVB exposure and cell viability was determined by MTT assay. UVB decreased cell viability by 60% compared to that of untreated control cells, whereas RA increased cell viability in a concentration-dependent manner, and this increase was significantly higher than that in the single treatment groups. Further, UVB increased the levels of DNA lesions such as cyclobutane pyrimidine dimer (CPD) and 8-hydroxyguanine (8-OHdG). Conversely, RA decreased both CPD and 8-OHdG levels in a concentration-dependent manner. UVB exposure also increased phosphorylation of ataxia-telangiectasia mutated (ATM) protein kinase and p53 and subsequently increased the levels of GADD45α, p21, and matrix metalloproteinases (MMPs)-3, -9, and -13. Additionally, UVB exposure decreased the level of COL1A1. However, RA treatment decreased the levels of p-ATM, p-p53, GADD45α, p21, MMP-3, -9, and -13 and increased the level of COL1A1 in a concentration-dependent manner. These results suggest that RA reduces UVB-induced cytotoxicity and genotoxicity through up-regulation of DNA repair via the combined effects of Rg2 and astaxanthin.

Comparison of Percutaneous Endoscopic Lumbar Diskectomy and Open Lumbar Microdiskectomy for Recurrent Lumbar Disk Herniation.

OBJECTIVE: To establish the appropriateness of transforaminal percutaneous endoscopic lumbar diskectomy (PELD) by comparing it with open lumbar microdiskectomy (OLM) for surgical treatment of recurrent lumbar disk herniation. METHODS: We retrospectively analyzed 83 patients who underwent revision surgery (group A: PELD, 35 cases; group B: OLM, 48 cases) for recurrent lumbar disk herniation between March 2009 and April 2014. All of the patients were followed > 12 months. To evaluate outcomes, we checked the leg and back visual analog scale (VAS), Oswestry Disability Index (ODI), complications, and recurrence for all patients. RESULTS: The mean improvement of VAS for leg was 5.97 ± 0.98 for group A and 5.62 ± 1.42 for group B (p = 0.194). The mean improvement of VAS for back pain was 2.71 ± 1.30 for group A and 2.29 ± 1.41 for group B (p = 0.168). The mean improvement of ODI scores was 28.86 ± 3.93 for group A and 28.00 ± 4.22 for group B (p = 0.350). Total surgery-related complications were none for group A and nine for group B (p = 0.009). Group A had one subject with surgery-related neurologic symptoms and group B had four (p = 0.391). Recurrence occurred in two patients in group A and seven patients in group B (p = 0.291). CONCLUSION: PELD and OLM have favorable clinical outcomes in patients with recurrent lumbar disk herniation, and PELD results in fewer complications compared with OLM. Therefore, PELD may be a better alternative to OLM for patients with recurrent lumbar disk herniation.

Tranexamic acid inhibits melanogenesis by activating the autophagy system in cultured melanoma cells.

BACKGROUND: As interest in skin beauty increases, the development of new skin whitening agents has attracted substantial attention; however, the action mechanism of the agents developed so far remains largely unknown. Tranexamic acid (TXA) is commonly being used to reduce melanin synthesis in patients with melasma and also used as a raw material for functional whitening cosmetics, although its action mechanism is poorly understood. Autophagy has been well known to be essential for tissue homeostasis, adaptation to starvation, and removal of dysfunctional organelles or pathogens. Recent studies have shown that autophagy regulators might have prominent roles in the initial formation stage of the melanosome, a lysosome-related organelle synthesizing melanin pigments. However, there is still no direct evidence showing a relationship between the activation of the autophagy system and the melanogenesis. OBJECTIVE: To investigate whether TXA can inhibit melanogenesis through the activation of autophagy in a melanoma cell line. METHODS: B16-F1 melanoma cells were treated with TXA and the levels of autophagy- and melanogenesis-related proteins were determined by Western blottings. The direct effect of TXA-mediated autophagy activation on melanin production was further evaluated by transfecting the cells with 60 pmols of small interfering RNAs (siRNAs)-targeting the mechanistic target of rapamycin (mTOR) and the autophagy-related protein 5 (Atg5). RESULTS: The results of Western blottings showed that TXA enhanced the production of autophagy-related proteins such as mitogen-activated protein kinases (MAPKs), extracellular signal-regulated kinase (ERK)1/2, Beclin-1, Atg12, and light chain 3 (LC3) I-II, whereas it decreased the synthesis of the mTOR complex. Confocal microscopy clearly showed that TXA treatment resulted in the formation of autophagosomes in B16-F1 cells, as revealed by immunostaining with an anti-LC3 antibody. The production of melanogenesis-associated proteins, including microphthalmia-associated transcription factor (MITF), tyrosinase, and tyrosinase-related protein 1 and 2 (TRP1/2), were clearly downregulated by the treatments with TXA. These results suggest that TXA can mediate a decrease in melanin synthesis by alleviating the production of tyrosinase and TRP1/2, along with lowered MITF protein levels. Furthermore, after treatment with TXA, siRNAs- targeting to mTOR and Atg5 increased melanin synthesis by 20% and 40%, respectively, compared to that in non-transfected cells, in a dose-dependent manner. These results further confirmed that TXA can inhibit melanogenesis by activating the autophagy system. CONCLUSION: Collectively, the results demonstrate that TXA can reduce melanin synthesis in melanoma B16-F1 cells by activating the ERK signaling pathway and the autophagy system.

Percutaneous Full Endoscopic Treatment of Bertolotti Syndrome: A Report of Three Cases with Technical Note.

Background Bertolotti syndrome is characterized by an abnormal enlargement of the transverse process of the most caudal lumbar vertebra. Most of the time it is asymptomatic, but when it is symptomatic it is associated with low back pain, radiating leg pain, or both. There is no consensus regarding management of this pathology. Open to minimal invasive tubular resection techniques are described in the literature, but a full endoscopic resection technique has not yet been described. Endoscopic technique is a less invasive target-oriented iliolumbar ligament preserving technique. We report our percutaneous full endoscopic technique for the treatment of symptomatic Bertolotti syndrome. Method We have treated three symptomatic cases of Bertolotti syndrome with the percutaneous endoscopic technique. We review the feasibility of this technique and the outcomes. Result Full endoscopic resection of Bertolotti syndrome is feasible and safe with comparable good outcomes and the added benefits of minimal invasive surgery. Conclusion We believe this novel percutaneous full endoscopic technique will yield good results in the hands of expert endoscopic spine surgeons and can be an alternative treatment method in cases of symptomatic Bertolotti syndrome. To our knowledge this is the first report of percutaneous full endoscopic treatment of Bertolotti syndrome in the world.