Experimental Biology and Medicine

In recent years, the “Copper-2 Hypothesis” has been put forth in an attempt to explain the epidemic of Alzheimer’s disease (AD) in the Western world. According to this hypothesis, “free” copper (copper-2) in drinking water, dietary supplements, and meat is the chief cause of the increased incidence of AD in recent decades. In contrast to the US, copper plumbing for drinking water is not used in Switzerland and tap water is very low in copper. Other “risk” factors including dietary supplements and meat consumption are also lower in Switzerland than in the US. Yet, the incidence of AD is closely similar in the two countries. This contradicts the Copper-2 Hypothesis.

Chondrocyte mechanotransduction is not well understood, but recently, it has been proposed that mechanically activated ion channels such as transient receptor potential vanilloid 4 (TRPV4), Piezo1, and Piezo2 are of functional importance in chondrocyte mechanotransduction. The aim of this study was to distinguish the potential contributions of TRPV4, Piezo1, and Piezo2 in transducing different intensities of repetitive mechanical stimulus in chondrocytes. To study this, TRPV4-, Piezo1-, or Piezo2-specific siRNAs were transfected into cultured primary chondrocytes to knock down (KD) TRPV4, Piezo1, or Piezo2 expression, designated TRPV4-KD, Piezo1-KD, or Piezo2-KD cells. Then we used Flexcell® Tension System to apply cyclic tensile strains (CTS) of 3% to 18% at 0.5 Hz for 8 h to the knockdown and control siRNA-treated cells. Finally, using a Ca²⁺ imaging system, stretch-evoked intracellular Ca²⁺ ([Ca²⁺] _i ) influx in chondrocytes was examined to investigate the roles of TRPV4, Piezo1, and Piezo2 in Ca²⁺ signaling in response to different intensities of repetitive mechanical stretch stimulation. The characteristics of [Ca²⁺] _i in chondrocytes evoked by stretch stimulation were stretch intensity dependent when comparing unstretched cells. In addition, stretch-evoked [Ca²⁺] _i changes were significantly suppressed in TRPV4-KD, Piezo1-KD, or Piezo2-KD cells compared with control siRNA-treated cells, indicating that any channel essential for Ca²⁺ signaling induced by stretch stimulation in chondrocytes. Of note, they played different roles in calcium oscillation induced by different intensities of stretch stimulation. More specifically, TRPV4-mediated Ca²⁺ signaling played a central role in the response of chondrocytes to physiologic levels of strain (3% and 8% of strain), while Piezo2-mediated Ca²⁺ signaling played a central role in the response of chondrocytes to injurious levels of strain (18% of strain). These results provide a basis for further examination of mechanotransduction in cartilage and raise a possibility of therapeutically targeting Piezo2-mediated mechanotransduction for the treatment of cartilage disease induced by repetitive mechanical forces.

Smoking has become a major cause of chronic obstructive pulmonary disease through weakening of the respiratory mucus-ciliary transport system, impairing cough reflex sensitivity, and inducing inflammation. Recent researches have indicated that hydrogen sulfide is essential in the development of various lung diseases. However, the effect and mechanism of hydrogen sulfide on cigarette smoke-induced chronic obstructive pulmonary disease have not been reported. In this study, rats were treated with cigarette smoke to create a chronic obstructive pulmonary disease model followed by treatment with a low concentration of hydrogen sulfide. Pulmonary function, histopathological appearance, lung edema, permeability, airway remodeling indicators, oxidative products/antioxidases levels, inflammatory factors in lung, cell classification in bronchoalveolar lavage fluid were measured to examine the effect of hydrogen sulfide on chronic obstructive pulmonary disease model. The results showed that hydrogen sulfide effectively improved pulmonary function and reduced histopathological changes, lung edema, and permeability. Airway remodeling, oxidative stress, and inflammation were also reduced by hydrogen sulfide treatment. To understand the mechanisms, we measured the expression of TGF-β1, TGF-βIand TGF-βII receptors and Smad7 and phosphorylation of Smad2/Smad3. The results indicated that the TGF-β1 and Smad were activated in cigarette smoke-induced chronic obstructive pulmonary disease model, but inhibited by hydrogen sulfide. In conclusion, this study showed that hydrogen sulfide treatment alleviated cigarette smoke-induced chronic obstructive pulmonary disease through inhibition of the TGF-β1/Smad pathway.

Friedreich’s ataxia is an autosomal recessive disorder characterized by impaired mitochondrial function, resulting in oxidative stress. In this study, we aimed at evaluating whether tocotrienol, a phytonutrient that diffuses easily in tissues with saturated fatty layers, could complement the current treatment with idebenone, a quinone analogue with antioxidant properties. Five young Friedreich’s ataxia patients received a low-dose tocotrienol supplementation (5 mg/kg/day), while not discontinuing idebenone treatment. Several oxidative stress markers and biological parameters related to oxidative stress were evaluated at the time of initiation of treatment and 2 and 12 months post-treatment. Some oxidative stress-related parameters and some inflammation indices were altered in Friedreich’s ataxia patients taking idebenone alone and tended to be normal values following tocotrienol supplementation; likewise, a cardiac magnetic resonance study showed some improvement following one-year tocotrienol treatment. The pathway by which tocotrienol affects the Nrf2 modulation of hepcidin gene expression, a peptide involved in iron handling and in inflammatory responses, is viewed in the light of the disruption of the iron intracellular distribution and of the Nrf2 anergy characterizing Friedreich’s ataxia. This research provides a suitable model to analyze the efficacy of therapeutic strategies able to counteract the excess free radicals in Friedreich’s ataxia, and paves the way to long-term clinical studies.

Pseudomonas aeruginosa-mannose-sensitive hemagglutinin (PAM) is an inactivate P. aeruginosa with mannose-sensitive hemagglutinin. Recently, the anticancer properties of PAM against many cancers have been reported across a range of studies. However, the exact mechanism through which PAM prevents skin cancer remains unclear. The aim of this study is to show to what extent PAM could inhibit the dimethylbenzanthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA)-induced skin cancer. JB6 cells were treated by TPA so as to establish an in vitro model. The effects of PAM on proliferation of the cells were analyzed using cell counting kit-8 assays. Effects on epithelial–mesenchymal transition (EMT) were assayed by real-time PCR and Western blotting. A DMBA/TPA-induced skin cancer mouse model was also established. The results showed that TPA promoted EMT changes through the activation of the hedgehog (Hh) pathway, which was reversed by PAM. Moreover, PAM inhibited the cancer growth and Hh pathway in vivo. These data indicate that PAM may serve as a potential anticancer agent for the treatment of skin cancer.

Inflammation plays a crucial part in hyperglycemia-induced myocardial damage. Hydrogen sulfide has been found to possess multiple biological activities in previous studies. This study investigated whether hydrogen sulfide conferred cardiac protection against damage in a diabetic rat model by inhibiting nucleotide-binding oligomerization domain-like receptor protein (NLRP) 3 inflammasome activation. Male animals were assigned to control, streptozotocin, streptozotocin + sodium hydrosulfide, and streptozotocin + DL-propargylglycine groups. Animals in the three streptozotocin groups were administrated 55 mg/kg streptozotocin by intraperitoneal injection. Streptozotocin + sodium hydrosulfide and streptozotocin + propargylglycine groups were treated with sodium hydrosulfide (56 μmol/kg) and propargylglycine (40 mg/kg), respectively, for four weeks. Estimation of fasting blood glucose, heart-weight/body-weight, cardiac function, and histopathological analysis, and measurement of myocardial enzymes were done to evaluate the degree of cardiac injury. In order to investigate the redox changes, the levels of total antioxidant capacity, malondialdehyde and lipid peroxidation, and the activities of superoxide dismutase, catalase, and glutathione peroxidase were assessed; the protein expression levels of Thioredoxin and Thioredoxin-interacting protein were measured in myocardial tissue. In addition, inflammatory reactions were assessed by measuring the concentration levels of interleukin-6, tumor necrosis factor-α, interleukin-1β, and interleukin-18 in serum and the expression levels of NLRP3 inflammasome complex-associated proteins in cardiac tissue. In the heart, hyperglycemia significantly induced cardiac dysfunction and injury, redox perturbation, and aggravation of inflammatory reactions. However, except for fasting blood glucose, treatment with sodium hydrosulfide significantly ameliorated these alterations, whereas treatment with propargylglycine further aggravated these alterations. This study highlights the protective properties of hydrogen sulfide against hyperglycemia-induced cardiac injury, and its possible mechanism was shown to involve negative regulation of Thioredoxin-interacting protein-mediated NLRP3 inflammasome activation.

Alzheimer’s disease as a consequence of chronic brain inflammation mediated by infectious microbes including the oral microbiome continues to attract support. Taiwan’s National Insurance database was used to evaluate associations between dental health and Alzheimer’s disease; 209,112 new cases of Alzheimer’s disease were matched 1:4 with 836,448 dementia-free controls to test the hypothesis that better dental health would be associated with less occurrence of dementia. Ten year dental records and conditional logistic regression models were used to estimate the odds ratios associated with Alzheimer’s disease. Subgroup analyses compared vascular Alzheimer’s disease and sporadic Alzheimer’s disease. As the population aged, Alzheimer’s disease diagnoses were more frequent with a 10 fold upward inflection after 60. Nearly 56% of sporadic Alzheimer’s disease patients were women but less than 50% had vascular Alzheimer’s disease. Comorbidities were 10–20% higher in the Alzheimer’s disease patients than in controls, but stroke, chronic infection, and pneumonia were 40–45% more common in the vascular Alzheimer’s disease patients. Heart disease, hypertension, diabetes, stroke, peripheral artery disease, pneumonia, and herpetic disease (HSV) were all associated with higher odds of Alzheimer’s disease. HSV was not a factor in the vascular Alzheimer’s disease. Routine dental procedures tended to lower odds ratios. Root canals and extractions that restore oral homeostasis were associated with lower odds of dementia. However, when extractions exceeded four, the odds of Alzheimer’s disease rose. The fact that Alzheimer’s disease was not associated with periodontal procedures per se but with more frequent periodontal emergencies suggested again a chronic issue. Dental health costs suggest that good dental care was associated with lower odds of Alzheimer’s disease except for radiographic costs which were consistently associated with higher odds, independent of oral health. Common comorbid conditions were associated with higher odds of Alzheimer’s disease and oral health care was associated with lower odds, providing support for the hypothesis that the oral microbiome is a factor in the development of Alzheimer’s disease.

Hypertensive status induces modifications in the respiratory profile. Previous studies have indicated that hypertensive rats show increased respiratory-sympathetic coupling compared to normotensive rats. However, these effects and especially the mechanisms underlying such effects are not well known. Thus, we evaluated the influence of high blood pressure and autonomic dysfunction on a ventilatory pattern associated with lung injury and on the ubiquitin-proteasome system of the diaphragm muscle. Autonomic cardiovascular modulation (systolic BP variance and low-frequency band and pulse interval variance) and arterial blood gases patterns (pH, pO₂, HCO₃, SpO₂), can be changed by hypertension, as well exacerbated chemoreflex pressor response. We observed that the diaphragm muscle of SHR showed increase in type I cross-sectional fiber (16%) and reduction in type II cross-sectional fiber area (41%), increased activity of the ubiquitin-proteasome system and lipid peroxidation, with no differences between groups in the analysis of ubiquitinated proteins and misfolded proteins. Our results showed that hypertension induced functional compensatory/adverse alterations associated with diaphragm fiber type changes and protein degradation as well as changed autonomic control of circulation. In conclusion, we believe there is an adaptation in ventilatory pattern in regarding to prevent the development of fatigue and muscle weakness and improve ventilatory endurance.

Natural plasma anti-α-galactoside antibody (anti-Gal) reactivity was reported to vary inversely with the individual’s lipoprotein(a) [Lp(a)] size. Since MUC1 mucin over-expressed in tumors bear surrogate peptide ligands for anti-Gal, we examined if high anti-Gal reactivity in small size/high titer Lp(a) individuals correlated with lower incidence of breast cancer. Newer protocol for size determination revealed that Lp(a) in controls were significantly smaller than in breast cancer patients (P = 0.0023; n = 46 in either group). Activity per unit plasma volume and specific reactivity (reactivity per unit immunoglobulin) of anti-Gal were significantly lower in cancer patients (P = 0.0033). Specific reactivity lower than the mean of controls was a risk factor for breast cancer with odds ratio (OR) 3.2 (95% confidence interval [CI]: 1.368–7.557). Immunochemical staining using fluorescein isothiocyanate-labeled anti-Gal revealed absolute inactivity towards normal cells and strong recognition of cancer cells by the antibody. O-Glycosylation of MUC1, though more frequent than in normal cells, was incomplete in tumor cells as revealed by binding of the O-glycan-specific lectin jacalin, accounting for the access of anti-Gal to its peptide ligand in cancer MUC1. As tumor advanced and MUC1 with increasing affinity for anti-Gal was synthesized by the tumor, the specific reactivity of circulating anti-Gal also increased, apparently due to antigenic stimulation or affinity maturation by the proliferating MUC1, indicating that pre-cancer anti-Gal reactivity in patients should have been much lower than measured after detection of cancer and that lower reactivity of the antibody is a stronger risk factor for breast cancer than indicated by the OR above. Reactivity towards a given group of tumor MUC1 antigens increased in proportion to anti-Gal specific reactivity. Results suggested tumor-specific MUC1 as likely target for anti-Gal-mediated anti-cancer defense and offer infusion of small Lp(a) or high reactivity anti-Gal as possible immunopotentiation measures.