Intron retention in PI-PLC γ1 mRNA as a key mechanism affecting MMP expression in human primary fibroblast-like synovial cells.

The intron retention (IR) is a phenomenon utilized by cells to allow diverse fates at the same mRNA, leading to a different pattern of synthesis of the same protein. In this study, we analyzed the modulation of phosphoinositide-specific phospholipase C (PI-PLC) enzymes by Harpagophytum procumbens extract (HPE) in synoviocytes from joins of osteoarthritis (OA) patients. In some samples, the PI-PLC γ1 isoform mature mRNA showed the IR and, in these synoviocytes, the HPE treatment increased the phenomenon. Moreover, we highlighted that as a consequence of IR, a lower amount of PI-PLC γ1 was produced. The decrease of PI-PLC γ1 was associated with the decrease of metalloprotease-3 (MMP-3), and MMP-13, and ADAMTS-5 after HPE treatment. The altered expression of MMPs is a hallmark of the onset and progression of OA, thus substances able to decrease their expression are very desirable. The interesting outcomes of this study are that 35% of analyzed synovial tissues showed the IR phenomenon in the PI-PLC γ1 mRNA and that the HPE treatment increased this phenomenon. For the first time, we found that the decrease of PI-PLC γ1 protein in synoviocytes interferes with MMP production, thus affecting the pathways involved in the MMP expression. This finding was validated by the silencing of PI-PLC γ1 in synoviocytes where the IR phenomenon was not present. Our results shed new light on the biochemical mechanisms involved in the degrading enzyme production in the joint of OA patients, suggesting a new therapeutic target and highlighting the importance of personalized medicine.

Osteoarthritis: Insights into Diagnosis, Pathophysiology, Therapeutic Avenues, and the Potential of Natural Extracts.

Osteoarthritis (OA) stands as a prevalent and progressively debilitating clinical condition globally, impacting joint structures and leading to their gradual deterioration through inflammatory mechanisms. While both non-modifiable and modifiable factors contribute to its onset, numerous aspects of OA pathophysiology remain elusive despite considerable research strides. Presently, diagnosis heavily relies on clinician expertise and meticulous differential diagnosis to exclude other joint-affecting conditions. Therapeutic approaches for OA predominantly focus on patient education for self-management alongside tailored exercise regimens, often complemented by various pharmacological interventions primarily targeting pain alleviation. However, pharmacological treatments typically exhibit short-term efficacy and local and/or systemic side effects, with prosthetic surgery being the ultimate resolution in severe cases. Thus, exploring the potential integration or substitution of conventional drug therapies with natural compounds and extracts emerges as a promising frontier in enhancing OA management. These alternatives offer improved safety profiles and possess the potential to target specific dysregulated pathways implicated in OA pathogenesis, thereby presenting a holistic approach to address the condition's complexities.

The Nutraceuticals as Modern Key to Achieve Erythrocyte Oxidative Stress Fighting in Osteoarthritis.

Osteoarthritis (OA), the most common joint disease, shows an increasing prevalence in the aging population in industrialized countries. OA is characterized by low-grade chronic inflammation, which causes degeneration of all joint tissues, such as articular cartilage, subchondral bone, and synovial membrane, leading to pain and loss of functionality. Erythrocytes, the most abundant blood cells, have as their primary function oxygen transport, which induces reactive oxygen species (ROS) production. For this reason, the erythrocytes have several mechanisms to counteract ROS injuries, which cause damage to lipids and proteins of the cell membrane. Oxidative stress and inflammation are highly correlated and are both causes of joint disorders. In the synovial fluid and blood of osteoarthritis patients, erythrocyte antioxidant enzyme expression is decreased. To date, OA is a non-curable disease, treated mainly with non-steroidal anti-inflammatory drugs and corticosteroids for a prolonged period of time, which cause several side effects; thus, the search for natural remedies with anti-inflammatory and antioxidant activities is always ongoing. In this review, we analyze several manuscripts describing the effect of traditional remedies, such as Harpagophytum procumbens, Curcumin longa, and Boswellia serrata extracts, in the treatments of OA for their anti-inflammatory, analgesic, and antioxidant activity. The effects of such remedies have been studied both in in vitro and in vivo models, considering both joint cells and erythrocytes.

The Fight against Infection and Pain: Devil's Claw (Harpagophytum procumbens) a Rich Source of Anti-Inflammatory Activity: 2011-2022.

Harpagophytum procumbens subsp. procumbens (Burch.) DC. ex Meisn. (Sesame seed Family-Pedaliaceae) is a popular medicinal plant known as Devil's claw. It is predominantly distributed widely over southern Africa. Its impressive reputation is embedded in its traditional uses as an indigenous herbal plant for the treatment of menstrual problems, bitter tonic, inflammation febrifuge, syphilis or even loss of appetite. A number of bioactive compounds such as terpenoids, iridoid glycosides, glycosides, and acetylated phenolic compounds have been isolated. Harpagoside and harpagide, iridoid glycosides bioactive compounds have been reported in countless phytochemical studies as potential anti-inflammatory agents as well as pain relievers. In-depth studies have associated chronic inflammation with various diseases, such as Alzheimer's disease, obesity, rheumatoid arthritis, type 2 diabetes, cancer, and cardiovascular and pulmonary diseases. In addition, 60% of chronic disorder fatalities are due to chronic inflammatory diseases worldwide. Inflammation and pain-related disorders have attracted significant attention as leading causes of global health challenges. Articles published from 2011 to the present were obtained and reviewed in-depth to determine valuable data findings as well as knowledge gaps. Various globally recognized scientific search engines/databases including Scopus, PubMed, Google Scholar, Web of Science, and ScienceDirect were utilized to collect information and deliver evidence. Based on the literature results, there was a dramatic decrease in the number of studies conducted on the anti-inflammatory and analgesic activity of Devil's claw, thereby presenting a potential research gap. It is also evident that currently in vivo clinical studies are needed to validate the prior massive in vitro studies, therefore delivering an ideal anti-inflammatory and analgesic agent in the form of H. procumbens products.

DNA Barcode Authentication of Devil's Claw Herbal Dietary Supplements.

Devil's claw is the vernacular name for a genus of medicinal plants that occur in the Kalahari Desert and Namibia Steppes. The genus comprises two distinct species: Harpagophytum procumbens and H. zeyheri. Although the European pharmacopeia considers the species interchangeable, recent studies have demonstrated that H. procumbens and H. zeyheri are chemically distinct and should not be treated as the same species. Further, the sale of H. zeyheri as an herbal supplement is not legal in the United States. Four markers were tested for their ability to distinguish H. procumbens from H. zeyheri: rbcL, matK, nrITS2, and psbA-trnH. Of these, only psbA-trnH was successful. A novel DNA mini-barcode assay that produces a 178-base amplicon in Harpagophytum (specificity = 1.00 [95% confidence interval = 0.80-1.00]; sensitivity = 1.00 [95% confidence interval = 0.75-1.00]) was used to estimate mislabeling frequency in a sample of 23 devil's claw supplements purchased in the United States. PCR amplification failed in 13% of cases. Among the 20 fully-analyzable supplements: H. procumbens was not detected in 75%; 25% contained both H. procumbens and H. zeyheri; none contained only H. procumbens. We recommend this novel mini-barcode region as a standard method of quality control in the manufacture of devil's claw supplements.

Analysis of Effectiveness of a Supplement Combining Harpagophytum procumbens, Zingiber officinale and Bixa orellana in Healthy Recreational Runners with Self-Reported Knee Pain: A Pilot, Randomized, Triple-Blind, Placebo-Controlled Trial.

Recreational running (RR) is becoming a popular way to increase physical activity for improving health, together with a higher incidence of knee injuries. The aim was to analyze the effect of a four-week supplementation with a mixture of Harpagophytum procumbens, Zingiber officinale and Bixa orellana on males, middle-aged, RR with an undiagnosed knee discomfort. A randomized triple-blind placebo-control trial was conducted among male RR aged 40-60 years suffering from self-declared knee discomfort after training. Participants were assigned to supplementation (2 g/day in 6 doses; n = 13; intervention group (IG)) or matched placebo (n = 15; control group (CG)) for 4 weeks. At pre- and post-intervention, assessment of routine blood biomarkers, body composition, running biomechanics and body temperature was performed using standardized procedures. Machine learning (ML) techniques were used to classify whether subjects belonged to IG or CG. ML model was able to correctly classify individuals as IG or CG with a median accuracy of 0.857. Leg fat mass decreased significantly (p = 0.037) and a deeper reduction in knee thermograms was observed in IG (p < 0.05). Safety evaluation revealed no significant differences in the rest of parameters studied. Subjects belonging to IG or CG are clearly differentiated, pointing into an effect of the supplement of ameliorating inflammation.

Devil's claw (Harpagophytum procumbens) ameliorates the neurobehavioral changes and neurotoxicity in female rats exposed to arsenic.

Over 200 million people are exposed to arsenic worldwide in their daily lives. Arsenic is a toxic ubiquitous metalloid distributed in the ground water. From the last few decades it is obtaining considerable attention for its severe neurotoxic properties. In this study the neuroprotective efficacy of devil's claw (DCW), a potent antioxidant has been investigated against arsenic induced neurotoxicity in female rats. Neurotoxicity was established by oral administration of 13 mg/kg sodium arsenite. The animals were divided into five groups (n = 6) including normal control, disease/arsenic control, standard treatment (Apocynin, 10 mg/kg), DCW treatment I (DCW, 200 mg/kg) and DCW treatment II (DCW, 400 mg/kg). Exploratory, anxiety and motor coordination related behavior of the animals was assessed using hole-board, forced swimming, beam walk and elevated plus maze tests. Findings revealed that DCW treatment ameliorated anxiety and motor in-coordination in the rats compared to the arsenic control group. In addition, arsenic induced a significant oxidative stress in arsenic only treated group, whereas co-administration with DCW the oxidative stress was reduced prominently. Arsenic control group produced gliosis and nuclear pyknosis of the brain cells which were prominently suppressed with the treatment of DCW for 21 days. The activity of DCW was in correlation with the concentration of harpagoside in the serum estimated by the HPLC method, supports that harpagoside was the active constituent responsible for neuroprotective effect. Further findings are required to understand the molecular mechanisms involved in neuroprotective effect of harpagoside and DCW.

Multidirectional Pharma-Toxicological Study on Harpagophytum procumbens DC. ex Meisn.: An IBD-Focused Investigation.

In the present study, we investigated the water extract of Harpagophytum procumbens DC. ex Meisn. in an experimental model of inflammatory bowel diseases (IBDs). Additionally, a microbiological investigation was carried out to discriminate the efficacy against bacterial and fungal strains involved in IBDs. Finally, an untargeted proteomic analysis was conducted on more than one hundred colon proteins involved in tissue morphology and metabolism. The extract was effective in blunting the production of oxidative stress and inflammation, including serotonin, prostaglandins, cytokines, and transcription factors. Additionally, the extract inhibited the growth of Candida albicans and C. tropicalis. The extract was also able to exert a pro-homeostatic effect on the levels of a wide plethora of colon proteins, thus corroborating a protective effect. Conversely, the supraphysiological downregulation of cytoskeletal-related proteins involved in tissue morphology and antimicrobial barrier function suggests a warning in the use of food supplements containing H. procumbens extracts.

Toxicology studies of aqueous-alcohol extracts of Harpagophytum procumbens subsp. procumbens (Burch.) DC.Ex Meisn. (Pedaliaceae) in female and male rats.

BACKGROUND: A variety of medicinal products prepared from secondary tubers of Harpagophytum procumbens subsp. procumbens (Burch.) DC.ex Meisn. (Devil's Claw) and H. zeyheri are marketed in Africa, Europe, the United States, South America and elsewhere, where they are used for inflammatory and musculoskeletal conditions such as arthritis, lower back pain, rheumatism and neuralgia, etc. While clinical studies conducted over the last twenty years support the general safety of such products, infrequent gastrointestinal disturbances (diarrhea, nausea, vomiting, abdominal pain), headache, vertigo and hypersensitivity (allergic) reactions (rash, hives and face swelling) have been documented. Sex-related differences occur in the health conditions for which Devil's Claw products are used, so it is likely that usage is similarly sex-related and so might be side effects and potential toxicities. However toxicologic studies of Devil's Claw products have been conducted primarily with male animals. To address this deficit, we report toxicological studies in female and male rats of several H. procumbens (HP) aqueous-alcohol extracts chemically analyzed by UPLC-MS. METHODS: Female and male Sprague Dawley rats were studied for one and three months in groups differing by consumption of diets without and with HP extracts at a 7-10-fold human equivalent dose (HED). Sera were analyzed for blood chemistry, and heart, liver, lung, kidney, stomach, and small and large intestine tissues were examined for histopathology. Treatment group differences for blood chemistry were analyzed by ANOVA with Dunnett's test and significant group differences for endpoints with marginal distributional properties were verified using the Kruskal-Wallis test. Group differences for histopathology were tested using Chi Square analysis. RESULTS: Significant group by sex-related differences in blood chemistry were detected in both studies. Additionally, several sex-related differences occurred between the studies. However, significant histopathology effects associated with the consumption of the extracts were not detected. CONCLUSION: Toxicologic analysis of Devil's Claw extracts cause significant sex-related effects in blood chemistry. However, in our judgement, none of the observed effects suggest serious toxicity at these doses and durations. Subsequent toxicologic and clinical studies of H. procumbens and other medicines with similar properties should explore in greater detail the basis and consequences of potential sex-related effects.

Harpagophytum procumbens Extract Ameliorates Allodynia and Modulates Oxidative and Antioxidant Stress Pathways in a Rat Model of Spinal Cord Injury.

Spinal cord injury (SCI) is a deliberating disorder with impairments in locomotor deficits and incapacitating sensory abnormalities. Harpagophytum procumbens (Hp) is a botanical widely used for treating inflammation and pain related to various inflammatory and musculoskeletal conditions. Using a modified rodent contusion model of SCI, we explored the effects of this botanical on locomotor function and responses to mechanical stimuli, and examined possible neurochemical changes associated with SCI-induced allodynia. Following spinal cord contusion at T10 level, Hp (300 mg/kg, p.o.) or vehicle (water) was administered daily starting 24 h post-surgery, and behavioral measurements made every-other day until sacrifice (Day 21). Hp treatment markedly ameliorated the contusion-induced decrease in locomotor function and increased sensitivity to mechanical stimuli. Determination of Iba1 expression in spinal cord tissues indicated microglial infiltration starting 3 days post-injury. SCI results in increased levels of 4-hydroxynonenal, an oxidative stress product and proalgesic, which was diminished at 7 days by treatment with Hp. SCI also enhanced antioxidant heme oxygenase-1 (HO-1) expression. Concurrent studies of cultured murine BV-2 microglial cells revealed that Hp suppressed oxidative/nitrosative stress and inflammatory responses, including production of nitric oxide and reactive oxygen species, phosphorylation of cytosolic phospholipases A2, and upregulation of the antioxidative stress pathway involving the nuclear factor erythroid 2-related factor 2 and HO-1. These results support the use of Hp for management of allodynia by providing resilience against the neuroinflammation and pain associated with SCI and other neuropathological conditions.

Chemical composition of various plant extracts and their in vitro efficacy in control of Fasciola hepatica eggs / Composição química de vários extratos vegetais e sua eficácia in vitro no controle de ovos de Fasciola hepática

ABSTRACT: Fasciolosis has been diagnosed in cattle, goats, sheep and horses in southern and southeastern Brazil. Effective alternative treatments are the targets of study. One promising alternative is the use of plant extracts. The aim of this study was to perform phytochemical analysis of extracts of Eugenia uniflora L., Harpagophytum procumbens, Psidium guajava L. and Stryphnodendron adstringens, and to evaluate the in vitro efficacy of these extracts on ovicidal activity in Fasciola hepatica. Plant extracts were analyzed for phytochemical properties. F. hepatica eggs were collected directly from the gallbladders of animals diagnosed as positive for fasciolosis on post mortem examination. One hundred eggs were incubated with 3 ml of each extract at concentrations of 0.10%, 0.25% and 0.50%, albendazole 0.50% (positive control) or tap water (negative control). To determine anti larval efficacy of each plant extract, hatched eggs were counted and the averages were used. Phytochemical analysis revealed the presence of phenolic compounds, tannins and terpenes in most extracts. E. uniflora L. extract was 100% effective at 0.10%, H. procumbens was effective at 0.25% and P. guajava L. and S. adstringens extracts were 100% effective at all concentrations tested. Taken together, the data suggested that ovicidal activity in F. hepatica is due to the presence of these bioactive compounds.

RESUMO: A fasciolose tem sido diagnosticada em bovinos, caprinos, ovinos e equinos no sul e sudeste do Brasil, sendo que tratamentos alternativos mais eficazes são alvos de estudo. Umas das alternativas promissoras é o uso de extratos vegetais no controle dessa e outras enfermidades. O objetivo deste estudo foi realizar a análise fitoquímica dos extratos de Eugenia uniflora L., Harpagophytum procumbens, Psidium guajava L. e Stryphnodendron adstringens, além de avaliar a eficácia in vitro desses extratos na atividade ovicida em Fasciola hepatica. Os extratos vegetais foram obtidos e analisados para determinação fitoquímica. Ovos de F. hepatica foram coletados diretamente das vesículas biliares de animais diagnosticados como positivos para fasciolose no exame post mortem. Cem ovos foram incubados com três mililitros de cada extrato nas concentrações de 0,10%, 0,25% e 0,50%, de albendazol a 0,50% (controle positivo) e água de torneira (controle negativo). Para determinar a eficácia de cada extrato vegetal os ovos eclodidos foram contados, e a média utilizada para os cálculos de eficácia. A análise fitoquímica revelou a presença de compostos fenólicos, taninos e terpenos na maioria dos extratos. O extrato de E. uniflora L. apresentou eficácia de 100% na concentração de 0,10%, o de H. procumbens a 0,25% e os extratos de P. guajava L. e S. adstringens apresentaram 100% de eficácia em todas as concentrações testadas. Assim, sugere-se que a atividade ovicida em F. hepatica seja devido à presença desses compostos bioativos.

Quality Assessment of Commercial Spagyric Tinctures of Harpagophytum procumbens and Their Antioxidant Properties.

Preparations from the dried tubers of Harpagophytum procumbens (Burch.) DC ex Meisn, commonly known as devil's claw, are mainly used in modern medicine to relieve joint pain and inflammation in patients suffering from rheumatic and arthritic disorders. This paper describes for the first time the chemical profile of a commercial spagyric tincture (named 019) prepared from the roots of the plant. For comparison purposes, a commercial not-spagyric devil's claw tincture (NST) was also analyzed. Chemical investigation of the content of specialized metabolites in the three samples indicated that harpagoside was the main compound, followed by the two isomers acteoside and isoacteoside. Compositional consistence over time was obtained by the chemical fingerprinting of another spagyric tincture (named 014) from the same producer that was already expired according to the recommendation on the label of the product. The two spagyric preparations did not show significant compositional differences as revealed by HPLC and MS analyses, except for a decrease in harpagide content in the expired 014 tincture. Moreover, their antioxidant capacities as assessed by 2,2'-di-phenyl-1-picrylhydrazyl (DPPH) and 2.2'-azin-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) methods resulted in very similar IC50 values. The expired 014 tincture showed instead a lower IC50 value compared to the 019 and NST tinctures with the ferric reducing antioxidant potential (FRAP) assay, indicating a higher ferric-reducing antioxidant ability. Overall, these results indicated that the two preparations could generally maintain good stability and biological activity at least for the four years from the production to the expiration date.

Pharmacokinetics of harpagoside in horses after intragastric administration of a Devil's claw (Harpagophytum procumbens) extract.

Devil's claw is used for the treatment of inflammatory symptoms and degenerative disorders in horses since many years, but without the substantive pharmacokinetic data. The pharmacokinetic parameters of harpagoside, the main active constituent of Harpagophytum procumbens DC ex Meisn., were evaluated in equine plasma after administration of Harpagophytum extract FB 8858 in an open, single-dose, two-treatment, two-period, randomized cross-over design. Six horses received a single dose of Harpagophytum extract, corresponding to 5 mg/kg BM harpagoside, and after 7 days washout period, 10 mg/kg BM harpagoside via nasogastric tube. Plasma samples at certain time points (before and 0-24 hr after administration) were collected, cleaned up by solid-phase extraction, and harpagoside concentrations were determined by LC-MS/MS using apigenin-7-glucoside as internal standard. Plasma concentration-time data and relevant parameters were described by noncompartmental model through PKSolver software. Harpagoside could be detected up to 9 hr after administration. Cmax was found at 25.59 and 55.46 ng/ml, t1/2 at 2.53 and 2.32 hr, respectively, and tmax at 1 hr in both trials. AUC0-inf was 70.46 and 117.85 ng hr ml-1 , respectively. A proportional relationship between dose, Cmax and AUC was observed. Distribution (Vz /F) was 259.04 and 283.83 L/kg and clearance (CL/F) 70.96 and 84.86 L hr-1  kg-1 , respectively. Treatment of horses with Harpagophytum extract did not cause any clinically detectable side effects.

Bioactive Compounds and Extracts from Traditional Herbs and Their Potential Anti-Inflammatory Health Effects.

The inflammatory processes associated with several chronic illnesses like cardiovascular disease and cancer have been the focus of mechanistic studies of the pathogenicity of these diseases and of the use of different pharmacological and natural methods to prevent them. In this study we review the current evidence regarding the effectiveness of natural extracts from as-yet little-studied traditional botanical species in alleviating the inflammation process associated with several chronic diseases. Additionally, the intention is to expose the known pathways of action and the potential synergistic effects of the constituent compounds of the discussed extracts. It is noted that the here-studied extracts, which include black garlic rich in S-allylcystein, polyphenols from cat's claw (Uncaria tomentosa), devil's claw (Harpagophytum procumbens), camu-camu (Myrciaria dubia), and blackcurrant (Ribes nigrum), and citrus fruit extracts rich in hesperidin, have similar or greater effects than other, more extensively studied extracts such as tea and cocoa. The combined use of all of these extracts can give rise to synergetic effects with greater biological relevance at lower doses.

A rare cause of gastrointestinal obstruction. Drug poisoning as a surgical disorder?

A patient had intestinal obstruction due to a rare cause. The patient presented unusual signs and symptoms. Although we performed a thorough diagnostic workup (CT, ultrasound, radiography, endoscopy), only laparotomy revealed that a bezoar caused the intestinal obstruction. The bezoar consisted of a herbal preparation, which was mentioned by the patient twice as a possible cause of his symptoms. All in all, the patient was right.

Use of Innovative (Micro)Extraction Techniques to Characterise Harpagophytum procumbens Root and its Commercial Food Supplements.

INTRODUCTION: For the determination of harpagoside and the wide phenolic pattern in Harpagophytum procumbens root and its commercial food supplements, dispersive liquid-liquid microextraction (DLLME), ultrasound-assisted DLLME (UA-DLLME), and sugaring-out liquid-liquid extraction (SULLE) were tested and compared. OBJECTIVES: In order to optimise the extraction efficiency, DLLME and UA-DLLME were performed in different solvents (water and aqueous solutions of glucose, ß-cyclodextrin, (2-hydroxypropyl)-ß-cyclodextrin, sodium chloride, natural deep eutectic solvent, and ionic liquid). MATERIAL AND METHODS: The plant material was ground and sieved to obtain a uniform granulometry before extraction. Commercial food supplements, containing H. procumbens are commercially available in Italy. RESULTS: The most effective sodium chloride-aided-DLLME was then optimised and applied for analyses followed by HPLC-PDA. For comparison, microwave-assisted extraction was performed using the same solvents and the best results were obtained using 1% of ß-cyclodextrin or 15% of sodium chloride. CONCLUSION: All commercial samples respected the European Pharmacopoeia monograph for this plant material, showing a harpagoside content ≥ 1.2%. Copyright © 2017 John Wiley & Sons, Ltd.

Protective Effects Induced by Microwave-Assisted Aqueous Harpagophytum Extract on Rat Cortex Synaptosomes Challenged with Amyloid ß-Peptide.

Harpagophytum procumbens is a plant species that displays anti-inflammatory properties in multiple tissues. The iridoid glycosides arpagoside, harpagide, and procumbide appear to be the most therapeutically important constituents. In addition, harpagoside treatment exerted neuroprotective effects both in vitro and in vivo. Considering these findings, the aim of the present work is to explore the possible protective role of the previously described microwave-assisted aqueous extract of H. procumbens on rat hypothalamic (Hypo-E22) cells, and in rat cortex challenged with amyloid ß-peptide (1-40). In this context, we assayed the protective effects induced by H. procumbens by measuring the levels of malondialdehyde, 3-hydroxykynurenine (3-HK), brain-derived neurotrophic factor, and tumor necrosis factor-α, 3-HK. Finally, we evaluated the effects of H. procumbens treatment on cortex levels of dopamine, norepinephrine, and serotonin. H. procumbens extract was well tolerated by Hypo-E22 cells and upregulated brain-derived neurotrophic factor gene expression but down-regulated tumor necrosis factor-α gene expression. In addition, the extract reduced amyloid ß-peptide stimulation of malondialdehyde and 3-HK and blunted the decrease of dopamine, norepinephrine, and serotonin, in the cortex. In this context, our work supports further studies for the evaluation and confirmation of Harpagophytum in the management of the clinical symptoms related to Alzheimer's disease. Copyright © 2017 John Wiley & Sons, Ltd.

Optimization of Aqueous Extraction and Biological Activity of Harpagophytum procumbens Root on Ex Vivo Rat Colon Inflammatory Model.

Harpagophytum procumbens has a long story of use for the treatment of inflammatory diseases. Considering both the antiinflammatory effects of H. procumbens in multiple tissues and the stability of harpagoside in artificial intestinal fluid, the aim of the present study was to explore the possible protective role of a microwave-assisted aqueous Harpagophytum extract (1-1000 µg/mL) on mouse myoblast C2C12 and human colorectal adenocarcinoma HCT116 cell lines, and isolated rat colon specimens challenged with lipopolysaccharide (LPS), a validated ex vivo model of acute ulcerative colitis. In this context, we evaluated the effects on C2C12 and HCT116 viability, and on LPS-induced production of serotonin (5-HT), tumor necrosis factor (TNF)-α, prostaglandin (PG)E2 and 8-iso-prostaglandin (8-iso-PG)F2α . Harpagophytum extract was well tolerated by C2C12 cells, while reduced HCT116 colon cancer cell viability. On the other hand, Harpagophytum extract reduced H2 O2 -induced (1 mM) reactive oxygen species (ROS) production, in both cell lines, and inhibited LPS-induced colon production of PGE2 , 8-iso-PGF2α , 5-HT and TNFα. Concluding, we demonstrated the efficacy of a microwave-assisted Harpagophytum aqueous extract in modulating the inflammatory, oxidative stress and immune response in an experimental model of inflammatory bowel diseases (IBD), thus suggesting a rational use of Harpagophytum in the management and prevention of ulcerative colitis in humans. Copyright © 2017 John Wiley & Sons, Ltd.

Two parametric cell cycle analyses of plant cell suspension cultures with fragile, isolated nuclei to investigate heterogeneity in growth of batch cultivations.

Plant cell suspensions are frequently considered to be heterogeneous with respect to growth in terms of progression of the cells through the cell cycle and biomass accumulation. Thus, segregated data of fractions in different cycle phases during cultivation is needed to develop robust production processes. Bromodeoxyuridine (BrdU) incorporation and BrdU-antibodies or 5-ethynyl-2'-deoxyuridine (EdU) click-it chemistry are frequently used to acquire such information. However, their use requires centrifugation steps that cannot be readily applied to sensitive cells, particularly if nuclei have to be extracted from the protective cellular milieu and envelopes for DNA analysis. Therefore, we have established a BrdU-Hoechst stain quenching protocol for analyzing nuclei directly isolated from delicate plant cell suspension cultures. After adding BrdU to test Harpagophytum procumbens cell suspension cultures the cell cycle distribution could be adequately resolved using its incorporation for the following 72 h (after which BrdU slowed biomass accumulation). Despite this limitation, the protocol allows resolution of the cell cycle distribution of cultures that cannot be analyzed using commonly applied methods due to the cells' fragility. The presented protocol enabled analysis of cycling heterogeneities in H. procumbens batch cultivations, and thus should facilitate process control of secondary metabolite production from fragile plant in vitro cultures. Biotechnol. Bioeng. 2016;113: 1244-1250. © 2015 Wiley Periodicals, Inc.

A new iridoid diglucoside from Harpagophytum procumbens.

A new iridoid diglucoside has been isolated from an aqueous extract of Harpagophytum procumbens secondary roots, together with six known compounds. Its structure has been assigned as 6'-O-glucopyranosyl-8-O-trans-coumaroylharpagide by spectroscopic means.