St. Johns wort attenuates irinotecan-induced diarhea via down-regulation of intestinal pro-inflammatory cytokines and inhibition of intestinal epithelial apoptosis

Diarrhea is a common dose-limiting toxicity associated with cancer chemotherapy, in particular for drugs such as irinotecan (CPT-11), 5-fluouracil, oxaliplatin, capecitabine and raltitrexed. St. John's wort (Hypericum perforatum, SJW) has anti-inflammatory activity, and our preliminary study in the rat and a pilot study in cancer patients found that treatment of SJW alleviated irinotecan-induced diarrhea. In the present study, we investigated whether SJW modulated various pro-inflammatory cytokines including interleukins (IL-1β, IL-2, IL-6), interferon (IFN-γ) and tumor necrosis factor-α (TNF-α) and intestinal epithelium apoptosis in rats. The rats were treated with irinotecan at 60 mg/kg for 4 days in combination with oral SJW or SJW-free control vehicle at 400 mg/kg for 8 days. Diarrhea, tissue damage, body weight loss, various cytokines including IL-1β, IL-2, IL-6, IFN-γ and TNF-α and intestinal epithelial apoptosis were monitored over 11 days. Our studies demonstrated that combined SJW markedly reduced CPT-11-induced diarrhea and intestinal lesions. The production of pro-inflammatory cytokines such as IL-1β, IFN-γ and TNF-α was significantly up-regulated in intestine. In the mean time, combined SJW significantly suppressed the intestinal epithelial apoptosis induced by CPT-11 over days 5–11. In particular, combination of SJW significantly inhibited the expression of TNF-α mRNA in the intestine over days 5–11. In conclusion, inhibition of pro-inflammatory cytokines and intestinal epithelium apoptosis partly explained the protective effect of SJW against the intestinal toxicities induced by irinotecan. Further studies are warranted to explore the potential for STW as an agent in combination with chemotherapeutic drugs to lower their dose-limiting toxicities.

Monitoring of immune responses to a herbal immuno-modulator in patients with advanced colorectal cancer.

Many herbal medicines are widely used as immuno-modulators in Asian countries. Ganoderma lucidum (Lingzhi) is one of the most commonly used herbs in Asia and preclinical studies have established that the polysaccharide fractions of G. lucidum have potent immuno-modulating effects. However, clinical evidence for this is scanty. The present open-labeled study aimed to evaluate the effects of G. lucidum polysaccharides on selected immune functions in patients with advanced colorectal cancer. Forty-seven patients were enrolled and treated with oral G. lucidum at 5.4 g/day for 12 weeks. Selected immune parameters were monitored using various immunological methods throughout the study. In 41 assessable cancer patients, treatment with G. lucidum tended to increase mitogenic reactivity to phytohemagglutinin, counts of CD3, CD4, CD8 and CD56 lymphocytes, plasma concentrations of interleukin (IL)-2, IL-6 and interferon (IFN)-γ, and NK activity, whereas plasma concentrations of IL-1 and tumor necrosis factor (TNF)-α were decreased. For all of these parameters, no statistical significance was observed when a comparison was conducted between baseline and those values after a 12-week treatment with G. lucidum. The changes of IL-1 were correlated with those for IL-6, IFN-γ, CD3, CD4, CD8 and NK activity (p < 0.05) and IL-2 changes were correlated with those for IL-6, CD8 and NK activity. The results indicate that G. lucidum may have potential immuno-modulating effect in patients with advanced colorectal cancer. Further studies are needed to explore the benefits and safety of G. lucidum in cancer patients.

Pharmacokinetic mechanisms for reduced toxicity of irinotecan by coadministered Thalidomide

The clinical use of irinotecan (CPT-11) is hindered by dose-limiting diarrhea and myelosuppression. Recent clinical studies indicate that thalidomide, a known tumor necrosis factor-alpha inhibitor, ameliorated the toxicities induced by CPT-11. However, the mechanisms for this are unknown. This study aimed to investigate whether combination of thalidomide modulated the toxicities of CPT-11 using a rat model and the possible role of the altered pharmacokinetic component in the toxicity modulation using in vitro models. The toxicity model was constructed by treatment of healthy rats with CPT-11 at 60 mg/kg per day by intravenous (i.v.) injection. Body weight, acute and delayed-onset diarrhea, blood cell counts, and macroscopic and microscopic intestinal damages were monitored in rats treated with CPT-11 alone or combined therapy with thalidomide at 100 mg/kg administered by intraperitoneal (i.p.) injection. Single dose and 5-day multiple-dose studies were conducted in rats to examine the effects of concomitant thalidomide on the plasma pharmacokinetics of CPT-11 and its major metabolites SN-38 and SN-38 glucuronide (SN-38G). The effect of CPT-11 on thalidomide's pharmacokinetics was also checked. Rat liver microsomes and a rat hepatoma cell line, H4-II-E cells, were used to study the in vitro metabolic interactions between these two drugs. H4-II-E cells were also used to investigate the effect of thalidomide and its hydrolytic products on the transport of CPT-11 and SN-38. In addition, the effect of thalidomide and its hydrolytic products on rat plasma protein binding of CPT-11 and SN-38 was examined. Administration of CPT-11 by i.v. for 4 consecutive days to rats induced significant body weight loss, decrease in neutrophil and lymphocyte counts, severe acute- and delayed-onset diarrhea, and intestinal damages. These toxicities were alleviated when CPT-11 was combined with thalidomide. In both single-dose and 5-day multiple-dose pharmacokinetic study, coadministered thalidomide significantly increased the area under the plasma concentration-time curve (AUC) of CPT-11, but the AUC and elimination half-life (t(1/2)) of SN-38 were significantly decreased. However, CPT-11 did not significantly alter the pharmacokinetics of thalidomide. Thalidomide at 25 and 250 microM and its hydrolytic products at a total concentration of 10 microM had no significant effect on the plasma protein binding of CPT-11 and SN-38, except for that thalidomide at 250 microM caused a significant increase in the unbound fraction (f(u)) of CPT-11 by 6.7% (P < 0.05). The hydrolytic products of thalidomide (total concentration of 10 microM), but not thalidomide, significantly decreased CPT-11 hydrolysis by 16% in rat liver microsomes (P < 0.01). The formation of both SN-38 and SN-38G from CPT-11, SN-38 glucuronidation, or intracellular accumulation of both CPT-11 and SN-38 in H4-II-E cells followed Michaelis-Menten kinetics with the one-binding site model being the best fit for the kinetic data. Coincubation or 2-hr preincubation of thalidomide at 25 microM and 250 microM and its hydrolytic products at 10 microM did not show any significant effects on CPT-11 hydrolysis and SN-38 glucuronidation. However, preincubation of H4-II-E cells with thalidomide (250 microM), its hydrolytic products (total concentration of 10 microM), or phthaloyl glutamic acid (one major thalidomide hydrolytic product, 10 microM) significantly increased the intracellular accumulation of SN-38, but not CPT-11 (P < 0.01). The dose-limiting toxicities of CPT-11 were alleviated by combination with thalidomide in rats and the pharmacokinetic modulation by thalidomide may partially explain its antagonizing effects on the toxicities of CPT-11. The hydrolytic products of thalidomide, instead of the parental drug, modulated the hepatic hydrolysis of CPT-11 and intracellular accumulation of SN-38, probably contributing to the altered plasma pharmacokinetics of CPT-11 and SN-38. Further studies are needed to explore the role of both pharmacokinetics and pharmacodynamic components in the protective effect of thalidomide against the toxicities of CPT-11.

Novel agents that potentially inhibit irinotecan-induced diarrhea

Irinotecan (CPT-11, 7-ethyl-10-[4-(1-piperidino)-1-piperidino] carbonyloxycamptothecin) has exhibited clinical activities against a broad spectrum of carcinomas by inhibiting DNA topoisomerase I (Topo I). However, severe and unpredictable dosing-limiting toxicities (mainly myelosuppression and severe diarrhea) hinder its clinical use. The latter consists of early and late-onset diarrhea, occurring within 24 hr or ≥ 24 hr after CPT-11 administration, respectively. This review highlights novel agents potentially inhibiting CPT-11-induced diarrhea, which are designed and tested under guidance of disposition pathways and potential toxicity mechanisms. Early-onset diarrhea is observed immediately after CPT-11 infusion and probably due to the inhibition of acetylcholinesterase activity, which can be eliminated by administration of atropine. Lateonset diarrhea appears to be associated with intestinal exposure to SN-38 (7-ethyl-10-hydroxycamptothecin), the major active metabolite of CPT-11, which may bind to Topo I and induce apoptosis of intestinal epithelia, leading to the disturbance in the absorptive and secretory functions of mucosa. CPT-11 and SN-38 may also stimulate the production of pro-inflammatory cytokines and prostaglandins (PGs), thus inducing the secretion of Na⁺ and Cl^-. Early treatment of severe late-onset diarrhea with oral high-dose loperamide has decreased patient morbidity. Extensive studies have been conducted to identify other potential agents to ameliorate diarrhea in preclinical and clinical models. These include intestinal alkalizing agents, oral antibiotics, enzyme inducers, P-glycoprotein (PgP) inhibitors, cyclooxygenase-2 (COX-2) inhibitors, tumor necrosis factor-agr (TNF-α) inhibitors, or blockers of biliary excretion of SN-38. Further studies are needed to identify the molecular targets associated with CPT-11 toxicity and safe and effective agents for alleviating CPT-11-induced diarrhea.

Biliary intervention augments chemotactic reaction and aggravates cholestatic liver injury in rats

Background
Intervention of the biliary system is frequently done in patients with obstructive jaundice and is associated with significant morbidity and mortality. The pathogenesis is unknown.
Materials and methods
A rat model of bile duct ligation (BDL) for 2 weeks was established in which biliary intervention was feasible by injection of normal saline through an indwelling catheter in the bile ducts. Plasma levels of C-C chemokine MCP-1 and C-X-C chemokine MIP-2 were measured by using ELISA. Blood monocytes, Kupffer cells, and neutrophils in the liver were characterized with antibodies to ED1, ED2, and myeloperoxidase (MPO). Lipid peroxidation was measured by malondialdehyde contents and apoptosis by TUNEL stain of the liver.
Results
Biliary intervention resulted in an increase of plasma MCP-1 and MIP-2 proteins by 1 h, which declined to normal level by 3 h in both sham and BDL rats. The levels in BDL rats were significantly higher than in sham at most points. There was a transient increase of ED1- and ED2-positive cells and MPO-staining cells in sham rat liver by 1 h after intervention. ED2-positive cells increased significantly by 1 h, while ED1- and MPO-positive cells decreased, yet insignificantly after intervention in BDL rats. The cell counts in BDL were constantly higher than in sham. Malondialdehyde increased precipitously in BDL by 3 h and was significantly higher than in sham throughout the study period. Parenchymal liver injury, manifested by elevated ALT, as well as apoptosis and necrosis of liver cells, was significantly increased in BDL rats, but not in sham rats.
Conclusion
Biliary intervention augments chemokine expression, precipitates lipid peroxidation, and aggravates liver injury in cholestatic rats.

Effects of dietary omega-3 polyunsaturated fatty acids on brain gene expression

Polyunsaturated fatty acids (PUFA) are essential structural components of the central nervous system. Their role in controlling learning and memory has been well documented. A nutrigenomic approach with high-density microarrays was used to reveal brain gene-expression changes in response to different PUFA-enriched diets in rats. In aged rats fed throughout life with PUFA-enriched diets, genes with altered expressions included transthyretin, α-synuclein, and calmodulins, which play important roles in synaptic  plasticity and learning. The effect of perinatal omega-3 PUFA supply on gene expression later in life also was studied. Several genes showed similar changes in expression in rats fed omega-3-deficient diets in the perinatal period, regardless of whether they or their mothers were fed omega-3 PUFA-sufficient diets after giving birth. In this experiment, among the down-regulated genes were a kainate glutamate receptor and a DEAD-box polypeptide. Among the up-regulated genes were a chemokine-like factor, a tumor necrosis factor receptor, and cytochrome c. The possible involvement of the genes with altered expression attributable to different diets in different brain regions in young and aged rats and the possible mode of regulatory action of PUFA also are discussed. We conclude that PUFA-enriched diets lead to significant changes in expression of several genes in the central nervous tissue, and these effects appear to be mainly independent of their effects on membrane composition. The direct effects of PUFA on transcriptional modulators, the downstream developmentally and tissue-specifically activated elements might be one of the clues to understanding the beneficial effects of the omega-3 PUFA on the nervous system.

Conjugated linoleic acid suppresses myogenic gene expression in a model of human muscle cell inflammation

Proinflammatory cytokines, such as tumor necrosis factor (TNF)-{alpha}, contribute to muscle wasting in inflammatory disorders, where TNF{alpha} acts to regulate myogenic genes. Conjugated linoleic acid (CLA) has shown promise as an antiproliferative and antiinflammatory agent, leading to its potential as a therapeutic agent in muscle-wasting disorders. To evaluate the effect of CLA on myogenesis during inflammation, human primary muscle cells were grown in culture and exposed to varying concentrations of TNF{alpha} and the cis-9, trans-11 and trans-10, cis-12 CLA isomers. Expression of myogenic genes (Myf5, MyoD, myogenin, and myostatin) and the functional genes creatine kinase (CK) and myosin heavy chain (MHC IIx) were measured by real-time PCR. TNF{alpha} significantly downregulated MyoD and myogenin expression, whereas it increased Myf5 expression. These changes corresponded with a decrease in both CK and MHC IIx expression. Both isomers of CLA mimicked the inhibitory effect of TNF{alpha} treatment on MyoD and myogenin expression, whereas myostatin expression was diminished in the presence of both isomers of CLA either alone or in combination with TNF{alpha}. Both isomers of CLA decreased CK and MHC IIx expression. These findings demonstrate that TNF{alpha} can have specific regulatory effects on myogenic genes in primary human muscle cells. A postulated antiinflammatory role of CLA in myogenesis appears more complex, with an indication that CLA may have a negative effect on this process.

Mucosal vascular addressin cell adhesion molecule-1 is expressed outside the endothelial lineage on fibroblasts and melanoma cells

Mucosal vascular addressin cell adhesion molecule-1 (MAdCAM-1) is predominantly expressed on high endothelial venules in inflamed tissues where it assists with leucocyte extravasation. Here we report that MAdCAM-1 has the potential to be more widely expressed outside the endothelial cell lineage than previously appreciated. Thus, MAdCAM-1 RNA transcripts and cell-surface protein were expressed by NIH 3T3 fibroblasts following activation with tumour necrosis factor-alpha (TNF-alpha), and by freshly isolated and cultured primary mouse splenic and tail fibroblasts in the absence of TNF-alpha stimulation. They were constitutively expressed by B16F10 melanoma cells, and expression was enhanced by cell activation with TNF-alpha. Mucosal vascular addressin cell adhesion molecule-1 was expressed on the apical surface of isolated cells, but became predominantly localized to cell junctions in confluent cell monolayers, suggesting it may play a role in the homotypic aggregation of cells. Tumour necrosis factor-alpha enhanced the expression of a firefly luciferase reporter directed by the MAdCAM-1 promoter in NIH 3T3 and B16F10 cells. A DNA fragment extending from nt -1727 to -673 was sufficient to confer cell-type selective expression. Mucosal vascular addressin cell adhesion molecule-1 expressed by NIH 3T3 cells was biologically active, as it supported the adhesion of TK-1 T cells in an alpha4beta7-dependent fashion. The expression of MAdCAM-1 by fibroblasts, and melanomas suggests MAdCAM-1 may play a role in regulating host responses in the periphery, leucocyte transmigration across nonendothelial boundaries, or the homotypic interactions of some malignant melanomas.

The case | Allograft dysfunction in a patient with sickle cell disease

The article focuses on the study concerning the diagnosis of acute renal allograft in patient with sickle cell disease. The 33 year old underwent several surgeries with immediate graft function. The percutaneous renal biopsy of the patient shows focal cortical necrosis and acute tubular damage coherent with ischemia. It concludes that the allograft sickling is the primary cause of the acute renal dysfunction in patients with sickle cell disease.

Relationship between inflammatory cytokines and self-report measures of training overload

It has been purported that inflammatory cytokines may be responsible for the aetiology of overtraining. The aim of the present study was to investigate the relationship between self-reported measures of overtraining and inflammatory cytokines. Eight elite male rowers were monitored in their natural training environment for 8 weeks prior to the 2007 Rowing World Championships. During this period of intense endurance training, self-report measures of overtraining and inflammatory cytokines (Interleukin (IL)-1β, IL-6, IL-8, IL-10, IL-12p70, and Tumor Necrosis Factor (TNF)-) were assessed fortnightly. Consistent with previous findings, proinflammatory cytokines IL-1β and TNF- were significantly associated (p ≤ 0.05) with measures of depressed mood, sleep disturbances, and stress. Similarly, IL-6 was significantly associated (p ≤ 0.01) with measures of depressed mood, sleep disturbances, and fatigue. These results are consistent with previous hypotheses describing how overtraining may be caused by excessive cytokine release, and lend further support for a cytokine hypothesis of overtraining.

Impact of training on changes in perceived stress and cytokine production

Data on training of competitive athletes and the inflammatory response, and, more specifically, the utility of psychological inventories to monitor this response in regards to overreaching is limited. The aim of the present study was to investigate the association of the Perceived Stress Scale (PSS) and inflammatory markers in elite rowers. Eight rowers (males n = 4; females n = 4) were monitored over an 8-week training period, comprising 12 sessions each week and training an average 3.11 h·d-1. Training volume was periodized weekly while intensity was maintained throughout the study. Perceived stress was measured weekly pretraining, and capillary blood samples (500 μL) were taken post-training. Significant associations between perceived stress and cytokines interleukin-6 (p < .05) and tumor necrosis factor alpha (p < .05) were observed. While further investigation of the role of cytokines in the overtraining process is required, these data provide preliminary support for an association between perceived stress and the inflammatory responses to training.

A gene expression signature for insulin resistance

Insulin resistance is a heterogeneous disorder caused by a range of genetic and environmental factors, and we hypothesize that its aetiology varies considerably between individuals. This heterogeneity provides significant challenges to the development of effective therapeutic regimes for long-term management of type 2 diabetes. We describe a novel strategy, using large-scale gene expression profiling, to develop a Gene Expression Signature (GES) that reflects the overall state of insulin resistance in cells and patients. The GES was developed from 3T3-L1 adipocytes that were made ‘insulin resistant’ by treatment with tumour necrosis factor-alpha (TNFα) and then reversed with aspirin and troglitazone (‘re-sensitized’). The GES consisted of five genes whose expression levels best discriminated between the insulin resistant and insulin re-sensitized states. We then used this GES to screen a compound library for agents that affected the GES genes in 3T3- L1 adipocytes in a way that most closely resembled the changes seen when insulin resistance was successfully reversed using aspirin and troglitazone. This screen identified both known and new insulin sensitizing compounds including non-steroidal anti inflammatory agents, β-adrenergic antagonists, beta-lactams and sodium channel blockers. We tested the biological relevance of this GES in participants in the San Antonio Family Heart Study (n = 1,240) and showed that patients with the lowest GES scores were more insulin resistant (according to HOMA_IR and fasting plasma insulin levels, P < 0.001). These findings show that GES technology can be used for both the discovery of insulin sensitizing compounds and the characterization of patients into subtypes of insulin resistance according to GES scores, opening the possibility of developing a personalized medicine approach to type 2 diabetes.

Unintentional needlestick injuries in livestock production : a case series and review

Livestock producers and their employees sometimes experience unintentional needlestick injury (NSI) while vaccinating or injecting medications into animals. There is little published regarding the medical complications that can develop from this occupational exposure. The objectives of this study were to (1) perform a retrospective review of animal-related NSIs treated at a tertiary medical center of a rural state; and (2) review the risks of NSI and measures to decrease their occurrence. Medical records of patients with NSI related to animal injection were identified from the University of Iowa Hospitals and Clinics database from 2002 to 2008 and reviewed. Nine patients received medical care for NSI that occurred while vaccinating farm animals. Most common NSI site was the nondominant hand and most occurred while attempting to inject the animal. Soft tissue infection was common and all nine received oral and/or intravenous antibiotics. Two thirds required hospital admission. Three required surgery and one had a bedside incision and drainage procedure. One patient had a serious inflammatory reaction with necrosis in the leg due to the oil adjuvant in the animal vaccine. Another case had a probable mycetoma with osteomyelitis and soft tissue infection due to the bacteria Streptomyces, which is a NSI complication not previously reported. Although medical complications from farm-related NSIs do not appear to be common, this case series illustrates how these injuries can be debilitating, costly, and lead to loss of work time and productivity. Producers and employees who inject livestock need to be aware of the risks and utilize measures to decrease unintentional NSI.

Oxidative stress : emerging mitochondrial and cellular themes and variations in neuronal injury

Oxidative stress plays a central role in neuronal injury and cell death in acute and chronic pathological conditions. The cellular responses to oxidative stress embrace changes in mitochondria and other organelles, notably endoplasmic reticulum, and can lead to a number of cell death paradigms, which cover a spectrum from apoptosis to necrosis and include autophagy. In Alzheimer's disease, and other pathologies including Parkinson's disease, protein aggregation provides further cellular stresses that can initiate or feed into the pathways to cell death engendered by oxidative stress. Specific attention is paid here to mitochondrial dysfunction and programmed cell death, and the diverse modes of cell death mediated by mitochondria under oxidative stress. Novel insights into cellular responses to neuronal oxidative stress from a range of different stressors can be gained by detailed transcriptomics analyses. Such studies at the cellular level provide the key for understanding the molecular and cellular pathways whereby neurons respond to oxidative stress and undergo injury and death. These considerations underpin the development of detailed knowledge in more complex integrated systems, up to the intact human bearing the neuropathology, facilitating therapeutic advances.

Photothermally induced apoptosis of cancer cells using gold nanorods for low energy cancer therapy

Gold nanorods functionalised with transferrin were used for photothermally induced necrosis and apoptosis of cancer cells. It was observed that the laser energy required to induce cell apoptosis is significantly lower than that for cell necrosis, indicating that photothermally induced apoptosis can be used for medically safe laser cancer treatment.