Alpha-Synuclein Accumulation and Its Phosphorylation in the Enteric Nervous System of Patients Without Neurodegeneration: An Explorative Study.

Alpha-synuclein (α-Syn) is widely distributed and involved in the regulation of the nervous system. The phosphorylation of α-Syn at serine 129 (pSer129α-Syn) is known to be closely associated with α-Synucleinopathies, especially Parkinson's disease (PD). The present study aimed to explore the α-Syn accumulation and its phosphorylation in the enteric nervous system (ENS) in patients without neurodegeneration. Patients who underwent colorectal surgery for either malignant or benign tumors that were not suitable for endoscopic resection (n = 19) were recruited to obtain normal intestinal specimens, which were used to assess α-Syn immunoreactivity patterns using α-Syn and pSer129α-Syn antibodies. Furthermore, the sub-location of α-Syn in neurons was identified by α-Syn/neurofilament double staining. Semi-quantitative counting was used to evaluate the expression of α-Syn and pSer129α-Syn in the ENS. Positive staining of α-Syn was detected in all intestinal layers in patients with non-neurodegenerative diseases. There was no significant correlation between the distribution of α-Syn and age (p = 0.554) or tumor stage (p = 0.751). Positive staining for pSer129α-Syn was only observed in the submucosa and myenteric plexus layers. The accumulation of pSer129α-Syn increased with age. In addition, we found that the degenerative changes of the ENS were related to the degree of tumor malignancy (p = 0.022). The deposits of α-Syn were present in the ENS of patients with non-neurodegenerative disorders; particularly the age-dependent expression of pSer129α-Syn in the submucosa and myenteric plexus. The current findings of α-Syn immunostaining in the ENS under near non-pathological conditions weaken the basis of using α-Syn pathology as a suitable hallmark to diagnose α-Synucleinopathies including PD. However, our data provided unique perspectives to study gastrointestinal dysfunction in non-neurodegenerative disorders. These findings provide new evidence to elucidate the neuropathological characteristics and α-Syn pathology pattern of the ENS in non-neurodegenerative conditions.

Synchronization Theory-Based Analysis of Coupled Vibrations of Dual-Tube Coriolis Mass Flowmeters.

Certain nonlinear influences are found in dual-tube Coriolis mass flowmeters (CMFs). According to experimentation, a nonlinearity dominated by frequency-doubling signals can be observed in the measuring signal. In general, such nonlinear effects are simplified as linear systems or neglected through processing. In this paper, a simplified model has been constructed for dual-beam CMFs based on the theory of nonlinear dynamics, with the spring-damper system as the medium for the dual-beam coupled vibrations. Next, the dynamics differential equation of the coupled vibrations is set up on the basis of the Lagrangian equation. Furthermore, numerical solutions are obtained using the Runge-Kutta fourth-order method. The study then fits discrete points of the numerical solutions, which are converted into the frequency domain to observe the existence of frequency-doubling signal components. Our findings show that frequency-doubling components exist in the spectrogram, proving that these nonlinear influences are a result of the motions of coupled vibrations. In this study, non-linear frequency-doubling signal sources are qualitatively analyzed to formulate a theoretical basis for CMFs design.

Pea leafminer Liriomyza huidobrensis (Diptera: Agromyzidae) uses vibrational duets for efficient sexual communication.

The pea leafminer (Liriomyza huidobrensis) is a notorious pest of vegetables and ornamental plants worldwide. Despite a large number of studies on its biology and ecology, the courtship behavior and sexual communication of this species remain unclear. Here, we studied vibrational communication in the sexual interaction of the pea leafminer. On host plant leaves, females and males behaviorally displayed the bobbing-quivering alternation, which finally led to copulation. Moreover, records of laser vibrometry revealed three-signal duets underlying the behavioral alternation. Sexually mature males spontaneously emitted calls (MCs) to initiate the duets. The females rapidly responded to MCs by emitting replies (FRs) that are longer in duration. The FRs further triggered male replies (MRs) in their search for potential partners. Leafminer-produced vibrational signals convey efficient information to partners and generate pair formation on stretched substrates, such as plant leaves and nylon mesh, but cannot elicit responses on dense substrates, such as glass and plastic. Vibrational playbacks of both MCs and FRs can elicit replies in females and males, respectively. This study completely characterizes substrate-borne vibrational duets in a dipteran insect. The discovery of vibrational sex signals in the pea leafminer provides new insights for the development of novel approaches to control the pest and its relative species.

miR-130b participates in wear particle-induced inflammation and osteolysis via FOXF2/NF-κB pathway.

OBJECTIVE: To reveal other miR-130b-mediated signaling pathway in the involvement of wear particle-induced inflammation and osteolysis. MATERIALS AND METHODS: Particle-induced osteolysis (PIO) mice model was established. Secretion levels of TNF-α, IL-1ß, IL-6, and IL-10 were measured by ELISA. miR-130b and forkhead box F2 (FOXF2) mRNA were detected by qRT-PCR. Protein levels of FOXF2, phosphorylation-p65 (p-p65), and p-IκB were observed by Western blot. Luciferase reporter assay was performed to confirm the regulation of miR-130b on FOXF2. RESULTS: Compared with normal mice, secretion levels of TNF-α, IL-1ß, and IL-6 in PIO mice were significantly up-regulated and IL-10 was significantly down-regulated; miR-130b and p-p65 expressions were up-regulated and FOXF2 expression was down-regulated. In addition, the trends of miR-130b, FOXF2, and p-p65 expressions in Co-Cr-Mo treated Raw264.7 cells were the same as that in PIO mice. After transfection with miR-130b inhibitor, secretion levels of TNF-α, IL-1ß, and IL-6 in Raw264.7 cells were significantly decreased and secretion level of IL-10 was significantly increased. We also proved FOXF2 was a target of miR-130b, and FOXF2 siRNA increased secretion levels of TNF-α, IL-1ß, and IL-6 and decreased secretion level of IL-10. Finally, we found nuclear factor-kappa B (NF-κB) inhibitor BAY 11-7082 further decreased secretion levels of TNF-α, IL-1ß, and IL-6 and increased IL-10 level. CONCLUSION: The role of miR-130b/FOXF2/NF-κB pathway in PIO was firstly revealed, which provided new targets for the treatment of periprosthetic osteolysis.

MicroRNA-130b Promotes Wear Particle-Induced Osteolysis via Downregulating Frizzled-Related Protein (FRZB).

Periprosthetic osteolysis induced by wear particles can lead to aseptic loosening, one main reason of arthroplasty failure. However, the role of microRNA-130b (miR-130b) in particle-induced osteolysis (PIO) has not been explored yet. In this study, PIO models were established in C57BL/J6 mice via the implantation of Co-Cr-Mo alloy particles, and evaluated by detecting tartrate-resistant acid phosphatase (TRAP) activity and bone resorption in the calvaria. Mouse preosteoblast MC3T3-E1 cells were cultured to receive particle stimulation in vitro. Real time PCR and western blotting were performed to determine the expression levels of miR-130b and frizzled-related protein (FRZB), one potential target of miR-130b. Results showed upregulated miR-130b and downregulated FRZB in both PIO mice with remarkable osteolysis and particle-treated MC3T3-E1 cells showing inhibited proliferation and differentiation assayed by bromodeoxy urodine (BrdU) incorporation and alkaline phosphatase (ALP) activity respectively. Functional studies were conducted by transfection of miR-130b inhibitor in vitro or the injections of miR-130b inhibitor or small interfering RNA (siRNA) targeting FRZB in vivo. Interestingly, particle-induced inhibition on cell proliferation, differentiation and FRZB expression were all reversed by miR-130b silence. Luciferase report assays demonstrated that miR-130b indeed negatively regulated FRZB expression by targeting, while FRZB could reverse the opposed effect of miR-130b silence on PIO development. Therefore, the upregulated miR-130b in PIO models could act as one key regulator of PIO development, partly due to its negative regulation on FRZB.

Abrasive Endoprosthetic Wear Particles Inhibit IFN-γ Secretion in Human Monocytes Via Upregulating TNF-α-Induced miR-29b.

The adverse biological responses to prostheses wear particles commonly led to the failure of total hip arthroplasty. Among the released cytokines, interferon-γ (IFN-γ) has been found to be a critical functional factor during osteoclast differentiation. However, the molecular mechanism underlying the regulation of IFN-γ in wear particles-induced cells still needs to be determined. Four kinds of abrasive endoprosthetic wear particle were used to treat THP-1 cells, including polymethylmethacrylate (PMMA), zirconiumoxide (ZrO2), commercially pure titanium (cpTi), and titanium alloy (Ti-6Al-7Nb), with a concentration of 0.01, 0.05, 0.1, or 0.2 mg/ml for 48 h. The expression of IFN-γ and miR-29b was detected by real-time RT-PCR or ELISA. Luciferase reporter assay was performed to determine the regulation of miR-29b on IFN-γ. The effect of miR-29b inhibitor on the expression of wear particle-induced IFN-γ was detected. The expression of miR-29b was examined in THP-1 cells treated with tumor necrosis factor-alpha (TNF-α). The expression of IFN-γ was downregulated and the level of miR-29b was increased in THP-1 cells pretreated with wear particles. IFN-γ was a target of miR-29b. Wear particles inhibited the expression of IFN-γ through miR-29b. The expression of miR-29b was significantly reduced in THP-1 cells treated with TNF-α neutralizing antibody and particles comparing to that in the cells treated with particles alone. Wear particles inhibit the IFN-γ secretion in human monocytes, which was associated with the upregulating TNF-α-induced miR-29b.

MicroRNA MiR-130b promotes wear particle-induced osteolysis via down regulating frizzled-related protein (FRZB).

Periprosthetic osteolysis induced by wear particles can lead to aseptic loosening, one main reason of arthroplasty failure. However, the role of microRNA-130b (miR-130b) in particle-induced osteolysis (PIO) has not been explored yet. In this study, PIO models were established in C57BL/J6 mice via the implantation of Co-Cr-Mo alloy particles, and evaluated by detecting tartrate-resistant acid phosphatase (TRAP) activity and bone resorption in the calvaria. Mouse preosteoblast MC3T3-E1 cells were cultured to receive particle stimulation in vitro. Real time PCR and western blotting were performed to determine the expression levels of miR-130b and frizzled-related protein (FRZB), one potential target of miR-130b. Results showed upregulated miR-130b and downregulated FRZB in both PIO mice with remarkable osteolysis and particle-treated MC3T3-E1 cells showing inhibited proliferation and differentiation assayed by bromodeoxy urodine (BrdU) incorporation and alkaline phosphatase (ALP) activity respectively. Functional studies were conducted by transfection of miR-130b inhibitor in vitro or the injections of miR-130b inhibitor or small interfering RNA (siRNA) targeting FRZB in vivo. Interestingly, particle-induced inhibition on cell proliferation, differentiation and FRZB expression were all reversed by miR-130b silence. Luciferase report assays demonstrated that miR-130b indeed negatively regulated FRZB expression by targeting, while FRZB could reverse the opposed effect of miR-130b silence on PIO development. Therefore, the upregulated miR-130b in PIO models could act as one key regulator of PIO development, partly due to its negative regulation on FRZB.

[Protection and mechanism of shenqi compound for diabetic angiopathy model rats].

OBJECTIVE: To investigate the protective effect and mechanism of Shenqi Compound on diabetic angiopathy modeled rats. METHODS: Totally 18 SD rats were randomized into 3 groups, i.e., the normal control group, the diabetic mellitus (DM) group, and Shenqi Compound group, 6 in each group. The DM rat model was established by feeding high-fat diet (to induce hyperlipidemia) +intraperitoneal injection of small dose streptozotocin (STZ). Shenqi Compound was given to rats in the Shenqi Compound group at the daily dose of 2 g/kg. Equal volume of normal saline was given to rats in the model group and the normal control group by gastrogavage. All treatment was lasted for 12 weeks. Then 2-D and ultrasonic integrated backscatter technique were used to evaluate structural and functional changes of abdominal aorta in the progression of diabetic macroangiopathy. The fibrosis degree of the aorta vessel and myocardium capillaries were observed by using HE and Masson trichrome staining. The tension of the aortic vascular ring was determined. The transforming growth factor beta (TGF-beta) mRNA expression was detected by real time PCR (RT-PCR). The protein expression of TGF-beta, collagen I, collagen III, connective tissue growth factor (CTGF), and phosphorylation P38 MAPK were detected by Western blot. RESULTS: Compared with the normal control group, abdominal aortic systolic inner diameter, diastolic inner diameter, Peterson elastic modulus, stiffness index, and backscatter integral significantly increased; the rangeability of integral backscatter and the extension coefficient of cross section significantly decreased in the DM group (all P < 0.05). After 12 weeks aforesaid indices were obviously improved in the Shenqi Compound group (P < 0.05). Results of HE and Masson staining showed that the fibrosis degree of the aorta vessel and myocardium capillaries was obviously alleviated in rats of the Shenqi Compound group (P < 0.05). Results of the aortic vascular ring tension showed that acetylcholine induced vasodilatation and maximum diastolic percent were obviously elevated in the Shenqi Compound group (P < 0.05). Compared with the normal control group, the mRNA expression of TGF-beta, and the protein expression of TGF-beta, collagen I, and collagen III, and phosphorylation of P38 MAPK all significantly increased in the DM group (P < 0.05). Compared with the DM group, the mRNA expression of TGF-beta, and the protein expression of TGF-beta, collagen I, and collagen III, and phosphorylation of P38 MAPK all decreased (P < 0.05). CONCLUSIONS: Shenqi Compound could effectively improve the arterial function in diabetic marcoangiopathy and microvascular dysfunction. The mechanism might be due to the down-regulating the expression of TGF-beta, and further suppressing the phosphorylation of P38 MAPK, reducing the synthesis of collagen I and collagen III, therefore, ameliorating arterial and myocardial interstitial fibrosis.

A rabbit model to study regression of ventricular hypertrophy.

BACKGROUND: The purpose of this study is to develop and characterise a reproducible rabbit model of LVH regression following pressure-induced myocardial hypertrophy. MATERIALS AND METHODS: Without endotracheal intubation and mechanical ventilation, a median sternotomy was performed. The median incision was made exactly along the midline of the sternum. Ascending aortic diameter was reduced 50% above the aortic valve, which lead to an approximate 75% reduction in crosssectional area. The sham-operated rabbits underwent the same procedures without actual ligation of the aorta. The animals of early DB group and late DB group were subjected to the second operation for removing the ligation eight weeks or 16 weeks after the initial banding surgery, respectively. Echocardiography, haemodynamic assessment, histology, and electron microscopy were used to assess functional and structural aspects of myocardial hypertrophy at each time point. RESULTS: The pressure overload resulted in robust LVH assessed by echocardiography, haemodynamic assessment, LV weight/body weight ratios, histology, and electron microscopy. Ascending aortic debanding completely or partially reversed LVH. The degree of LVH regression was association with the duration of pressure overload, proved by the fact that restoration of LV structure and function was complete in animals subjected to eight weeks of aortic banding but incomplete in animals subjected to 16 weeks of aortic banding. The animals did not experience severe mechanical ventilatory impairment. CONCLUSIONS: These results demonstrate an efficient and reproducible method of promoting LVH regression in rabbits without endotracheal intubation and mechanical ventilation.

Ultrasonic frogs show extraordinary sex differences in auditory frequency sensitivity.

Acoustic communication has an important role in the reproductive behaviour of anurans. Although males of the concave-eared frog (Odorrana tormota) have shown an ultrasonic communication capacity adapted to the intense, predominately low-frequency ambient noise from local streams, whether the females communicate with ultrasound remains unclear. Here we present evidence that females exhibit no ultrasonic sensitivity. Acoustic playback experiments show that the calls from male evoke phonotaxis and vocal responses from gravid females, whereas the ultrasonic components (frequencies above 20 kHz) of the calls do not elicit any phonotaxis or vocalization in the females. Electrophysiological recordings from the auditory midbrain reveal an upper frequency limit at 16 kHz in females. Laser Doppler vibrometer measurements show that the velocity amplitude of the tympanic membranes peaks at 5 kHz in females and at ∼7 kHz in males. The auditory sex differences in O. tormota imply that ultrasonic hearing has evolved only in male anurans.

Orexigenic effect of cocaine- and amphetamine-regulated transcript (CART) after injection into hypothalamic nuclei in streptozotocin-diabetic rats.

1. The aim of the present study was to investigate the orexigenic effect of cocaine- and amphetamine-regulated transcript (CART) peptide on feeding regulation following its injection into discrete nuclei of the hypothalamus. 2. Male Sprague-Dawley diabetic rats were injected with 0.06 or 0.2 nmol CART (55-102) or an equal volume of saline into various hypothalamic areas and food intake was then measured 1, 2, 4, 8 and 24 h after injection. Changes in hypothalamic CART mRNA expression in response to dietary intervention (2 weeks feeding of a high-fat diet) were assessed using quantitative real-time reverse transcription-polymerase chain reaction. Possible interactions between neuropeptide Y (NPY), agouti-related protein (AGRP), α-melanocyte-stimulating hormone (MSH) and corticotropin-releasing hormone (CRH) were evaluated in an in vitro hypothalamic explant system. Neuropeptide immunoreactivities (IR) were determined using radioimmunoassays (RIAs). 3. At 0.2 nmol, CART (55-102) significantly increased feeding in fasted diabetic rats after injection into the dorsomedial hypothalamic nucleus and arcuate nucleus (ARC). Injection of 0.2 nmol CART (55-102) into the ARC of satiated diabetic rats also increased food intake that was similar in both magnitude and time-course to the response seen in fasted diabetic rats. Food intake in diabetic rats on a high-fat diet was clearly increased after injection of 0.2 nmol CART (55-102) into the ARC, as was CART mRNA expression. Incubation of hypothalamic explants with 0.4, 4 and 40 nmol/L CART (55-102) for 45 min significantly increased NPY IR, whereas exposure of explants to 4 nmol/L CART (55-102) increased AGRP IR and CRH IR. None of the concentrations of CART (55-102) tested had any effect on α-MSH IR. 4. Together, these data provide further evidence that hypothalamic CART has an orexigenic effect, which, in the ARC, may stimulate the release of hypothalamic orexigenic neuropeptides.

Animal model of ascending aortic stenosis in rabbits without endotracheal intubation.

BACKGROUND AND AIM: Experimental animal studies of left ventricular hypertrophy (LVH) usually involve endotracheal intubation, which is associated with a risk of serious tracheal injury and other significant negative sequelae. We developed an animal model of pressure overload hypertrophy caused by constriction of the ascending aorta in rabbits that does not require endotracheal intubation. METHODS: New Zealand White rabbits of either sex (1.94 +/- 0.12 kg) were randomly assigned to the aortic banding (AB) group (n = 9) and the sham-operated group (n = 8). The sternum was carefully incised along the midline to avoid injury to the parietal pleura. Then, the intervention group underwent AB with three to zero Prolene proximal to the innominate artery without endotracheal intubation. To investigate the effects of the surgical procedure on physiological parameters, echocardiography, histology, and electron microscopy were performed to assess functional and structural hypertrophy at various time intervals. RESULTS: Banding of the ascending aorta created the expected increases in both aortic velocity and the pressure gradient between proximal and distal aorta coarctation. The pressure overload resulted in a robust LVH assessed by transthoracic echocardiography and histology. The animals did not experience severe mechanical ventilatory impairment. CONCLUSION: We developed a safe, efficient, and reproducible method of producing LVH in rabbits without the need for endotracheal intubation and mechanical ventilation. Ultimately, this model will facilitate focused study of the mechanisms involved with LVH progression.