Pogostemon cablin essential oil affects anxiety- and depressive-like behaviors and the gut microbiota in chronic unpredictable mild stress model rats.

The gut microbiota is thought to be an important factor that influences brain processes and behaviors through the gut-brain axis. Pogostemon cablin is used in traditional Chinese medicine (TCM) to treat gastrointestinal symptoms. Patchouli essential oil (PCO), the main active agent in P. cablin, is used in aromatherapy for stress relief. The aim of our study was to investigate the effects of orally administered PCO on anxiety- and depressive-like behaviors and the gut microbiota. We constructed a rat model of chronic unpredictable mild stress (CUMS) and explored the anxiolytic- and antidepressant-like effects of PCO using the open field test (OFT) and forced swim test (FST). Changes in the abundance of the gut microbiota, short-chain fatty acids (SCFAs), and other related molecules were assessed to determine the role of the gut microbiota. Our results showed that CUMS induced an anxiety-like phenotype in the OFT, which was reversed by PCO, and that PCO also significantly mitigated the depression-like behaviors caused by CUMS in the FST. Furthermore, we found that PCO increased the relative abundances of several probiotics, including Bacteroides and Blautia, and decreased the relative abundances of Ruminococcus_1 and Ruminococcus_2, which were increased by CUMS. Regarding SCFAs, the metabolites of the gut microbiota, PCO increased the concentration of propionic acid and decreased that of caproic acid. Finally, PCO restored the serotonin (5-hydroxytryptamine, 5-HT) level in the hippocampus, which had been decreased by CUMS. The results of this study suggested that PCO can improve stress-related anxiety- and depression-like behaviors and might exert its effects on the central nervous system through interactions with the gut microbiota.

Cyclic Glycine-Proline (cGP) Normalises Insulin-Like Growth Factor-1 (IGF-1) Function: Clinical Significance in the Ageing Brain and in Age-Related Neurological Conditions.

Insulin-like growth factor-1 (IGF-1) function declines with age and is associated with brain ageing and the progression of age-related neurological conditions. The reversible binding of IGF-1 to IGF binding protein (IGFBP)-3 regulates the amount of bioavailable, functional IGF-1 in circulation. Cyclic glycine-proline (cGP), a metabolite from the binding site of IGF-1, retains its affinity for IGFBP-3 and competes against IGF-1 for IGFBP-3 binding. Thus, cGP and IGFBP-3 collectively regulate the bioavailability of IGF-1. The molar ratio of cGP/IGF-1 represents the amount of bioavailable and functional IGF-1 in circulation. The cGP/IGF-1 molar ratio is low in patients with age-related conditions, including hypertension, stroke, and neurological disorders with cognitive impairment. Stroke patients with a higher cGP/IGF-1 molar ratio have more favourable clinical outcomes. The elderly with more cGP have better memory retention. An increase in the cGP/IGF-1 molar ratio with age is associated with normal cognition, whereas a decrease in this ratio with age is associated with dementia in Parkinson disease. In addition, cGP administration reduces systolic blood pressure, improves memory, and aids in stroke recovery. These clinical and experimental observations demonstrate the role of cGP in regulating IGF-1 function and its potential clinical applications in age-related brain diseases as a plasma biomarker for-and an intervention to improve-IGF-1 function.

Exploring ex vivo peptideolysis of thymopentin and lipid-based nanocarriers towards oral formulations.

The oral delivery of medicines is the most popular route of administration for patients. However, thymopentin (TP5) is only available in the market in forms for parenteral administration. In large part, this is because of extensive peptidolytic degradation in the gastrointestinal tract (GIT), which decreases the amount of TP5 available for absorption. This study aims to understand the extent of TP5 peptideolysis and determine effective inhibitors and suitable lipid-based nanocarriers to aid in the development of an effective oral delivery formulation. Enzymatic degradation kinetics of TP5 was investigated in the presence or absence of mucosal and luminal components extracted from various parts of the rat intestine, including the duodenum, jejunum, ileum, and colon. Inhibition of TP5 enzymatic peptidolysis was screened in the presence or absence of EDTA, trypsin and chymotrypsin inhibitors from soybean (SBTCI), and bestatin. TP5 with SBTCI was loaded into lipid-based nanocarriers, including microemulsions, niosomes and solid lipid nanoparticles. These TP5-loaded nanocarriers were investigated through characterization of morphology, particle size, zeta potential, entrapment efficacy (EE%), and ex vivo rat intestinal degradation studies to select a lead formulation for a future oral drug delivery study. The degradation kinetics of TP5 followed pseudo-first-order kinetics, and the biological metabolism of TP5 was displayed in the presence of luminal contents, indicating that TP5 is sensitive to luminal enzymes. Notably, a considerable decrease in TP5 peptidolysis was found in the presence of SBTCI, bestatin, and EDTA. TP5 and SBTCI were loaded into three lipid-based delivery systems, displaying superior protection under ex vivo intestinal luminal contents and mucosal homogenates for 6 h compared with the pure drug solution. These findings suggest that using select inhibitors and lipid-based nanocarriers can decrease peptide degradation and may improve oral bioavailability of TP5 following oral administration.

Administration of cyclic glycine-proline during infancy improves adult spatial memory, astrocyte plasticity, vascularization and GluR-1 expression in rats.

Cyclic glycine-proline (cGP) is a natural nutrient of breast milk and plays a role in regulating the function of insulin-like growth factor-1 (IGF-1). IGF-1 function is essential for post-natal brain development and adult cognitive function. We evaluated the effects of cGP on spatial memory and histological changes in the hippocampus of the adult rats following infancy administration. Infant rats were treated with either cGP or saline between post-natal days 8 and 22 via oral administration to lactating dams. The spatial memory was evaluated between post-natal days 70 and 75 using Morris water maze tests. The changes of capillaries, astrocytes, synaptophysin and glutamate receptor-1 were examined in the CA1 stratum radiatum of the hippocampus. Compared to saline-treated group, cGP-treated group showed higher path efficiency of entry and lower average heading errors to the platform zone. cGP-treated group also showed longer, larger and more astrocytic processes, more capillaries and higher glutamate receptor-1 expression. The rats made less average heading error to the platform zone have more capillaries, larger and longer astrocytic branches. Thus cGP treatment/supplementation during infancy moderately improved adulthood spatial memory. This long-lasting effect of cGP on memory could be mediated via promoting astrocytic plasticity, vascularization and glutamate trafficking. Therefore, cGP may have a role in regulating IGF-1 function during brain development.

The autocrine regulation of insulin-like growth factor-1 in human brain of Alzheimer's disease.

BACKGROUND: Insulin-like growth factor (IGF) binding protein (IGFBP)-3 and cyclic Glycine-Proline (cGP) regulate circulating IGF-1 function that is associated with cognition. The association between IGF-1 function and Alzheimer's disease (AD) remains inconclusive. This study evaluated the changes of IGFBPs and cGP, and their effects on the bioavailability and function of IGF-1 in human brain of AD cases. METHODS: Using biological and mathematic analysis we measured the concentrations of total, bound and unbound forms of IGF-1, IGFBPs and cGP in the inferior-frontal gyrus and middle-frontal gyrus of human AD (n = 15) and control cases (n = 15). The association between the changes of total concentration of these peptides and total protein concentration in brain tissues were also analyzed. RESULTS: The unbound bioavailable IGF-1 was lower whereas the bound cGP and IGFBP-3 were higher in AD than the control cases. Total protein that was lower in AD than control cases, was negatively associated with cGP concentration of control cases and with IGFBP-3 concentration of AD cases. CONCLUSIONS: The results provide direct evidence for IGF-1 deficiency in AD brain due to lower bioavailable IGF-1. The increase of bound IGFBP-3 impaired autocrine regulation. The increase of bound cGP is an autocrine response to improve the bioavailability and function of IGF-1 in AD brain. AVAILABILITY OF DATA AND MATERIAL: All data generated or analysed during this study are included in this published article. Additional datasets analysed during the current study available from the corresponding author on reasonable request.

Honokiol-Chlorambucil Co-Prodrugs Selectively Enhance the Killing Effect through STAT3 Binding on Lymphocytic Leukemia Cells In Vitro and In Vivo.

The broad-spectrum DNA alkylating therapeutic, chlorambucil (CBL), has limited safety and shows lower therapy effect because of a short half-life while used in the clinic. Therefore, it is very necessary to develop a more efficient and safer type of CBL derivate against tumors with selective targeting of cancer cells. In addition, the natural product of honokiol (HN), the novel potent chemo-preventive or therapeutic entity/carrier, can target the mitochondria of cancer cells through STAT3 to prevent cancer from spreading and metastasizing. In this study, we designed and synthesized the honokiol-chlorambucil (HN-CBL) co-prodrugs through carbonate ester linkage conjugating with the targeted delivery help of the HN skeleton in cancer cells. Biological evaluation indicated that HN-CBL can remarkably enhance the antiproliferation of human leukemic cell lines CCRF-CEM, Jurkat, U937, MV4-11, and K562. Furthermore, HN-CBL can also selectively inhibit the lymphocytic leukemia (LL) cell survival compared to those mononuclear cells derived from healthy donors (PBMCs), enhance mitochondrial activity in leukemia cells, and induce LL cell apoptosis. Molecular docking and western blot study showed that HN-CBL can also bind with the STAT3 protein at some hydrophobic residues and downregulate the phosphorylation level of STAT3-like HN. Significantly, HN-CBL could dramatically delay leukemia growth in vivo with no observable physiological toxicity. Thus, HN-CBL may provide a novel and effective targeting therapeutic against LL with fewer side effects.

Blood and brain protein levels of ubiquitin-conjugating enzyme E2K (UBE2K) are elevated in individuals with schizophrenia.

A number of recent studies have suggested the ubiquitin proteasome system (UPS) in schizophrenia is dysfunctional. The purpose of this study was to investigate UBE2K, a ubiquitin-conjugating (E2) enzyme within the UPS that has been associated with psychosis symptom severity, in the blood and brain of individuals with schizophrenia. Whole blood and erythrocytes from 128 (71 treatment-resistant schizophrenia, 57 healthy controls) individuals as well as frozen dorsolateral prefrontal cortex (DLPFC) and orbitofrontal cortex (OFC) post-mortem samples from 74 (37 schizophrenia, 37 controls) individuals were obtained. UBE2K gene expression was assayed in whole blood and DLPFC samples, whereas protein levels were assayed in erythrocytes and OFC samples. Elevated levels of UBE2K mRNA were observed in whole blood of individuals with schizophrenia (p = 0.03) but not in the DLPFC, while protein levels were raised in erythrocytes and the OFC (p < 0.001 and p = 0.002 respectively). Findings were not better explained by age, smoking, clozapine plasma levels or duration of illness. Although blood and brain samples were derived from independent samples, our findings suggest peripheral protein levels of UBE2K may serve as a surrogate of brain levels and further supports the notion of UPS dysfunction in schizophrenia. Future studies to determine the pathophysiological effects of elevated UBE2K protein levels in the brain of those with schizophrenia are warranted.

Elevated ubiquitinated proteins in brain and blood of individuals with schizophrenia.

Dysregulation of the ubiquitin proteasome system (UPS) has been linked to schizophrenia but it is not clear if this dysregulation is detectable in both brain and blood. We examined free mono-ubiquitin, ubiquitinated proteins, catalytic ubiquitination, and proteasome activities in frozen postmortem OFC tissue from 76 (38 schizophrenia, 38 control) matched individuals, as well as erythrocytes from 181 living participants, who comprised 30 individuals with recent onset schizophrenia (mean illness duration = 1 year), 63 individuals with 'treatment-resistant' schizophrenia (mean illness duration = 17 years), and 88 age-matched participants without major psychiatric illness. Ubiquitinated protein levels were elevated in postmortem OFC in schizophrenia compared to controls (p = <0.001, AUC = 74.2%). Similarly, individuals with 'treatment-resistant' schizophrenia had higher levels of ubiquitinated proteins in erythrocytes compared to those with recent onset schizophrenia (p < 0.001, AUC = 65.5%) and controls (p < 0.001, AUC = 69.4%). The results could not be better explained by changes in proteasome activity, demographic, medication, or tissue factors. Our results suggest that ubiquitinated protein formation may be abnormal in both the brain and erythrocytes of those with schizophrenia, particularly in the later stages or specific sub-groups of the illness. A derangement in protein ubiquitination may be linked to pathogenesis or neurotoxicity in schizophrenia, and its manifestation in the blood may have prognostic utility.

Development and Characterization of High-Throughput EST-Based SSR Markers for Pogostemon cablin Using Transcriptome Sequencing.

Simple sequence repeats (SSRs) or microsatellite markers derived from expressed sequence tags (ESTs) are routinely used for molecular assisted-selection breeding, comparative genomic analysis, and genetic diversity studies. In this study, we investigated 54,546 ESTs for the identification and development of SSR markers in Pogostemon cablin (Patchouli). In total, 1219 SSRs were identified from 1144 SSR-containing ESTs. Trinucleotides (80.8%) were the most abundant SSRs, followed by di- (10.8%), mono- (7.1%), and hexa-nucleotides (1.3%). The top six motifs were CCG/CGG (15.3%), AAG/CTT (15.0%), ACC/GGT (13.5%), AGG/CCT (12.4%), ATC/ATG (9.9%), and AG/CT (9.8%). On the basis of these SSR-containing ESTs, a total of 192 primer pairs were randomly designed and used for polymorphism analysis in 38 accessions collected from different geographical regions of Guangdong, China. Of the SSR markers, 45 were polymorphic and had allele variations from two to four. Furthermore, a transferability analysis of these primer pairs revealed a 10⁻40% cross-species transferability in 10 related species. This report is the first comprehensive study on the development and analysis of a large set of SSR markers in P. cablin. These markers have the potential to be used in quantitative trait loci mapping, genetic diversity studies, and the fingerprinting of cultivars of P. cablin.

Antidepressant-like effect of saponins extracted from Chaihu-jia-longgu-muli-tang and its possible mechanism.

In this study, we investigated the antidepressant-like effect of saponins (SCLM) extracted from a traditional Chinese medicine, Chaihu-jia-longgu-muli-tang (CLM), in mice and rats using the tail suspension test (TST) and forced swimming test (FST). Subchronic administration of 100 and 200 mg/kg (p.o.) SCLM for 7 days reduced immobility time in the TST and FST in mice and also decreased immobility time at 70 and 140 mg/kg (p.o.) in the FST in rats. The results also showed that the anti-immobility activity of SCLM in these two tests is dose-dependent, without accompanying significant effects on locomotor activity. In addition, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and lactic dehydrogenase (LDH) assays showed that 25, 50 and 100 microg/ml SCLM or 10 microM fluoxetine (FLU), protected PC12 cells from the lesion induced by 10 microM corticosterone (Cort) treatment for 48 h. In the fura-2/AM (acetoxymethyl ester) labeling assay, 50 and 100 microg/ml SCLM, 10 microM FLU attenuated the intracellular Ca2+ overloading induced by 200 microM Cort treatment for 48 h in PC12 cells. Using RT-PCR, the mRNA level of nerve growth factor (NGF) was also detected. Treatment with SCLM (50, 100 microg/ml) for 48 h elevated the NGF mRNA expression in PC12 cells. In summary, these results suggest that SCLM possesses an antidepressant-like activity in behavioral models that might be mediated via the cytoprotective action shown in PC12 cells.