Grb2 Y160F mutant mimics the wild-type monomeric state dynamics and the monomer-dimer equilibrium.

The Growth factor receptor-bound protein 2 (Grb2) participates in early signaling complexes and regulates tyrosine kinase-mediated signal transduction through a monomer-dimer equilibrium. Grb2 dimeric state inhibits signal transduction whereas the monomer promotes signaling downstream. Since Grb2 dimer KD is ∼0.8 µM, studies focused on the monomer are still challenging and require mutations or interaction with phosphotyrosine peptides. However, these mutants were never characterized considering their effects on protein structure and dynamics in solution. Here, we present the biophysical characterization of Grb2Y160F, the first Grb2 mutant to induce protein monomerization without disrupting its native behavior in solution due to net charge modifications or interaction with peptides. We also identified that Grb2Y160F exists in a monomer-dimer equilibrium. Grb2Y160F ability to dimerize implies that different dimerization interfaces might regulate signaling pathways in distinct ways and raises an important question about the role of the Y160 residue in other dimerization interfaces.

The real-world safety profile of tirzepatide: pharmacovigilance analysis of the FDA Adverse Event Reporting System (FAERS) database.

PURPOSE: Randomized controlled trials with tirzepatide (TZP) displayed unprecedented glucose and body weight lowering efficacy in individuals with type 2 diabetes and/or obesity and a safety profile similar to that of glucagon-like peptide-1 receptor agonists (GLP-1RA), mainly characterized by gastrointestinal (GI) adverse events (AE). Concerns on diabetic retinopathy, pancreato-biliary disorders, and medullary thyroid cancer were also addressed. We aimed to investigate whether the same safety issues emerged from the FDA Adverse Event Reporting System (FAERS) post-marketing surveillance database. METHODS: OpenVigil 2.1-MedDRA-v24 and AERSMine (data 2004Q1-2023Q3) were used to query the FAERS database. Reports of GI AE, diabetic retinopathy, pancreato-biliary disorders, and medullary thyroid cancer were investigated. The analysis was then filtered for age, gender, and designation as primary suspect. AE occurrence with TZP was compared to insulin, sodium-glucose cotransporter-2 inhibitors, metformin, and GLP-1RA. RESULTS: Disproportionate reporting of GI [i.e., nausea (ROR 4.01, 95% CI 3.85-4.19)] and pancreato-biliary disorders [i.e., pancreatitis (ROR 3.63, 95% CI 3.15-4.19)], diabetic retinopathy (ROR 4.14, 95% CI 2.34-7.30), and medullary thyroid cancer (ROR 13.67, 95% CI 4.35-42.96) was detected. TZP exhibited a similar risk of GI AE and medullary thyroid cancer and a lower risk of most pancreato-biliary AE and diabetic retinopathy vs. GLP-1RA. CONCLUSIONS: TZP was associated with an increased risk of specific AE. However, its safety profile was similar to that of GLP-1RA, without increased risk of pancreato-biliary AE, diabetic retinopathy, and medullary thyroid cancer.

Pharmacological modulation of adaptive thermogenesis: new clues for obesity management?

BACKGROUND: Adaptive thermogenesis represents the main mechanism through which the body generates heat in response to external stimuli, a phenomenon that includes shivering and non-shivering thermogenesis. The non-shivering thermogenesis is mainly exploited by adipose tissue characterized by a brown aspect, which specializes in energy dissipation. A decreased amount of brown adipose tissue has been observed in ageing and chronic illnesses such as obesity, a worldwide health problem characterized by dysfunctional adipose tissue expansion and associated cardiometabolic complications. In the last decades, the discovery of a trans-differentiation mechanism ("browning") within white adipose tissue depots, leading to the generation of brown-like cells, allowed to explore new natural and synthetic compounds able to favour this process and thus enhance thermogenesis with the aim of counteracting obesity. Based on recent findings, brown adipose tissue-activating agents could represent another option in addition to appetite inhibitors and inhibitors of nutrient absorption for obesity treatment. PURPOSE: This review investigates the main molecules involved in the physiological (e.g. incretin hormones) and pharmacological (e.g. ß3-adrenergic receptors agonists, thyroid receptor agonists, farnesoid X receptor agonists, glucagon-like peptide-1, and glucagon receptor agonists) modulation of adaptive thermogenesis and the signalling mechanisms involved.

New insights into the treatment of non-healing diabetic foot ulcers.

Diabetic foot ulcers (DFUs) are one of the most serious and devastating complication of diabetes mellitus, affecting about 15% of diabetic patients. This review describes the innovative treatment options currently available in the treatment of non-healing DFUs. The use of Platelet-Rich-Plasma (PRP) is a safe and valid approach in the treatment of DFUs. However, the methods used to obtain and prepare autologous PRP vary between the studies, thus further evidences are eagerly awaited. Adipose tissue-derived mesenchymal stem cells (ADSCs) are a promising tool in the treatment of DFUs, but additional largescale and long-term follow-up clinical trials are needed. Bone marrow mesenchymal stem cells (BM-MSCs) transplantation, on the other hand, revealed effective in reducing incidents and improving the quality of life of patients with amputations. Autologous Peripheral Blood Mononuclear Cells (A-PBMNCs) showed a good efficacy in the treatment of diabetic patients with CLI, but further RCTs are awaited to best investigate this new therapeutic approach. Photobiomodulation (PBM) therapy revealed effective in the treatment of DFUs in two RCTs, but a standardization of therapeutic protocols as well as level-I studies are needed.

Thyroid and shoulder diseases: the bases of a linked channel.

The association between thyroid disorders and musculoskeletal diseases has long been suspected, but it is still debated whether they have a role in the pathogenesis of shoulder diseases. In vivo and in vitro studies describe the role of thyroid hormones in bone, cartilage and tendon biology. Retrospective studies and case reports suggest that thyroid diseases should be considered as risk factors and hold prognostic value in some of the most common causes of shoulder pain. Thus, it is advisable to search for underlying thyroid disorders in these patients. The pathophysiologic mechanisms by which thyroid hormone imbalance affects the onset, progression and response to treatment of these diseases are yet to be thoroughly defined and demand further studies.

Active site mapping of Loxosceles phospholipases D: Biochemical and biological features.

Brown spider phospholipases D from Loxosceles venoms are among the most widely studied toxins since they induce dermonecrosis, triggering inflammatory responses, increase vascular permeability, cause hemolysis, and renal failure. The catalytic (H12 and H47) and metal-ion binding (E32 and D34) residues in Loxosceles intermedia phospholipase D (LiRecDT1) were mutated to understand their roles in the observed activities. All mutants were identified using whole venom serum antibodies and a specific antibody to wild-type LiRecDT1, they were also analyzed by circular dichroism (CD) and differential scanning calorimetry (DSC). The phospholipase D activities of H12A, H47A, H12A-H47A, E32, D34 and E32A-D34A, such as vascular permeability, dermonecrosis, and hemolytic effects were inhibited. The mutant Y228A was equally detrimental to biochemical and biological effects of phospholipase D, suggesting an essential role of this residue in substrate recognition and binding. On the other hand, the mutant C53A-C201A reduced the enzyme's ability to hydrolyze phospholipids and promote dermonecrosis, hemolytic, and vascular effects. These results provide the basis understanding the importance of specific residues in the observed activities and contribute to the design of synthetic and specific inhibitors for Brown spider venom phospholipases D.

Functional expression, monodispersity and conformational changes in the SBMV virus viral VPg on binding TFE.

Southern bean mosaic virus (SBMV) RNA purified from infected plants was used for cloning the viral genome-linked protein (VPg) and was subsequently expressed in Escherichia coli. Circular dichroism (CD), dynamic light scattering (DLS) and saturation transfer difference (STD) by nuclear magnetic resonance (NMR) measurements were employed to determine the degree of monodispersity and to investigate the conformational changes in the absence and presence of trifluoroethanol (TFE) which indicated increased helical content with increasing concentration of TFE. 8-Anilino-1-naphthalenesulfonic acid (ANS) was used as a probe to compare the unfolding regions of the protein before and after addition of TFE. The results indicated that although the TFE concentration influences VPg folding, it does not play a role in nucleotide binding and that the local solvent hydrophobicity causes significant conformational changes.

A method for positioning electrodes during surface EMG recordings in lower limb muscles.

PURPOSE: The aim of this work is to provide information about the degree of inter-subject uniformity of location of innervation zone (IZ) in 13 superficial muscles of the lower limb. The availability of such information will allow researchers to standardize and optimize their electrode positioning procedure and to obtain accurate and repeatable estimates of surface electromyography (sEMG) signal amplitude, spectral variables and muscle fiber conduction velocity. METHODS: Surface EMG signals from gluteus maximus, gluteus medius, tensor faciae latae, biceps femoris, semitendinosus, vastus medialis obliquus, vastus lateralis, rectus femoris, tibialis anterior, peroneus longus, soleus, gastrocnemius medialis and lateralis muscles of ten healthy male subjects aged between 25 and 34 years (average = 29.2 years, S.D. = 2.5 years) were recorded to assess individual IZ location and signal quality. RESULTS: Tensor faciae latae, biceps femoris, semitendinosus, vastus lateralis, gastrocnemius medialis and lateralis showed a high level of both signal quality and IZ location uniformity. In contrast, rectus femoris, gluteus medius and peroneus longus were found to show poor results for both indexes. Gluteus maximus, vastus medialis obliquus and tibialis anterior were found to show high signal quality but low IZ location uniformity. Finally, soleus muscle was found to show low signal quality but high IZ location uniformity. CONCLUSIONS: This study identifies optimal electrode sites for muscles in the lower extremity by providing a standard landmarking technique for the localization of the IZ of each muscle so that surface EMG electrodes can be properly positioned between the IZ and a tendon.

Surface EMG alterations induced by underwater recording.

BACKGROUND AND PURPOSE: This study aims to verify if amplitude and spectral characteristics of surface EMG signal are modified due to recording in a wet environment. METHODS: Isometric contractions of the biceps brachii muscle of ten subjects were performed in several different set-up combinations, both in dry (D) and in water from hydrotherapy pools (PW), with (PWM) or without moving the pool water and with (T) or without water-resistant adhesive taping. RESULTS: In PW condition the amplitude of the recorded signal is reduced to 5-10% of the corresponding signal recorded in D condition. In PWM the power spectrum is drastically reduced and altered by the water movement that introduces an increase of spectral power in the frequency range 0-20 Hz. The use of T modality allows to record signals with both amplitude and spectral frequencies comparable with those obtained in the D conditions. DISCUSSION AND CONCLUSION: This work demonstrates the need for water resistant taping when EMG signals are recorded in water. Signals recorded without such a protective film are strongly affected in their amplitude and frequency characteristics by the conductivity and the movement of water.

Bioelectrical impedance measures in different position and vs dual-energy X-ray absorptiometry (DXA).

BACKGROUND: Bioelectrical impedance analysis (BIA) is a safe, low-cost, non-invasive, rapid method for the assessment of body composition. It has therefore a great potential to be employed for epidemiological and clinical studies. However, many devices are available to estimate total body water (TBW), fat-free mass (FFM) and fat mass (FM) by bioelectrical impedance measurements. Moreover, bipedal devices allowing measurements in the only standing position are recently developed. They are easy and practical to use without operator, so a large diffusion can be forecasted in fields as sport and diet programs. Comparison of body composition estimation by a bipedal device with bioimpedance devices currently used, using dual-energy X-ray absorptiometry (DXA) as reference method. METHODS: The study was performed on 18 healthy women volunteers, age 32.0+/-10.7 years divided in two groups at different levels of body fatness. A Xitron 4000 impedance analyser, a BIA-101 RJL System, and the bipedal device Tanita were used for comparison. The measurements were performed in standing and supine position for Xitron and RJL devices. DXA measurements were performed with a total body scanner DPX, Lunar. RESULTS: FM and FFM were not statistically different when measured with Xitron and RJL in comparison with DXA, while these variables were significantly different between Tanita and DXA measurements. No significant difference were found between measurements in the supine and standing position with the Xitron and RJL system. CONCLUSIONS: Our results suggest that FM and FFM evaluated by bipedal device Tanita are significantly different from FM and FFM measured by DXA in both normal and obese population.

The role of NSAIDs in psoriatic arthritis: evidence from a controlled study with nimesulide.

OBJECTIVE: To define the optimal dose of nimesulide (NIM) for treating psoriatic arthritis. METHODS: Eighty patients entered a 4-week, double-dummy, randomised, controlled study. Each patient was allocated to one of the following treatment groups: NIM 100 mg/day, NIM 200 mg/day, NIM 400 mg/day, or placebo. Primary end points for arthritis assessment were the scores for tender and swollen joints, and the physician's and patient's global assessment of efficacy. RESULTS: 76/80 patients completed the study. NIM decreased the score for tender and swollen joints from baseline to the end swollen joints from baseline to the end of therapy (p < 0.05). Pain severity on a visual analogue scale (VAS) was reduced by NIM 200 mg (p = 0.03) or NIM 400 mg (p = 0.019) compared to placebo, as was morning stiffness (p = 0.038 and p = 0.008, respectively), but the trends with 100 mg were not statistically significant. The investigators and patients assessed the global efficacy of the NIM 200 and 400 mg/day groups as better than placebo or NIM 100 mg. Side effects were observed in 12 patients (15%) during treatment. They were mostly mild, only one patient withdrew from the study as a result, and the trend for a higher incidence with NIM was not statistically significant. The Psoriasis Area Severity Index (PASI) and the ESR did not show any significant changes. Patients in the placebo group took more paracetamol per day compared with those in the NIM groups (p = 0.007). CONCLUSIONS: Nimesulide 200 and 400 mg/day are effective and safe in psoriatic arthritis.

Oral type II collagen in the treatment of rheumatoid arthritis. A six-month double blind placebo-controlled study.

OBJECTIVE: To evaluate the efficacy of oral chicken type II collagen (CII) in the treatment of rheumatoid arthritis (RA). METHODS: Sixty patients with clinically active RA of long duration (mean 7.2 +/- 5.5 years) were treated for 6 months with oral chicken CII at 0.25 mg/day (n = 31) or with placebo (n = 29) in a double-blind randomized study. RESULTS: The response rate to treatment of the collagen-treated group, based on the ACR 20% criteria, was higher than that of the control group but this difference was not statistically significant at any time. Intention-to-treat (ITT) analysis did not show statistically significant improvement in any of the several secondary outcome measures over the 6 months of the study in the collagen-treated patients in comparison with the placebo-treated group. However, in 2 collagen-treated patients we observed a clinical remission according to the criteria of the American Rheumatism Association. CONCLUSION: Our study seems to show that the oral treatment of RA patients with chicken CII is ineffective and results in only small and inconsistent benefits. Furthermore, our results raise the possibility that in a sub-group of patients oral collagen administration, usually considered devoid of harmful effects, may actually induce disease flares.

Geometrical factors in surface EMG of the vastus medialis and lateralis muscles.

Surface EMG signals detected in dynamic conditions are affected by a number of artefacts. Among them geometrical factors play an important role. During movement the muscle slides with respect to the skin because of the variation of its length. Such a shift can considerably modify sEMG amplitude. The purpose of this work is to assess geometrical artefacts on sEMG during isometric contractions at different muscle lengths. The average rectified value (ARV) of 15 single differential signals was obtained by means of a linear array of 16 bar electrodes from the vastus medialis and lateralis muscles. The knee angle was changed from 75 degrees to 165 degrees in steps of 30 degrees and voluntary isometric contractions at a low, medium and high force level were performed for each angle. The ARV pattern was normalized with respect to the mean activity to compare signals from different joint angles. From the data collected it was possible to separate the geometrical changes from the changes due to different intensities of activation. In three out of five subjects, we found (within the resolution of our measures) a 1 cm shift for the vastus medialis muscle while no shift was observed for the other two subjects. For the vastus lateralis muscle a 1 cm shift was found in two out of four subjects. Such a shift produces the main contribution to geometrical artefacts. To avoid such artefacts the innervation zones should be located and the EMG electrodes should not be placed near them.

Amtolmetin guacyl versus piroxicam in patients with osteoarthritis.

The efficacy and tolerability of amtolmetin guacyl (AMG), a new non-steroidal anti-inflammatory drug, were compared with piroxicam, in patients with osteoarthritis. In a randomized double-blind study patients with arthritis (n = 99) received either 600 mg AMG on an empty stomach or 20 mg of piroxicam on a full stomach, once daily for 30 days. All clinical parameters improved significantly with both drugs; there were no significant differences between the two treatments. Tolerability, assessed by the patients, was significantly better in the AMG group. In the piroxicam group nine of 50 patients withdrew because of side-effects (gastrointestinal) compared with two of 49 (nausea and headache) in the AMG group. There were three cases of perforation, ulcer and bleeding in the piroxicam group but no serious side-effects with AMG. Total numbers of side-effects were similar in the two groups, but epigastric and abdominal pain were more frequent and more intense with piroxicam. AMG was as effective as piroxicam in controlling the symptoms of osteoarthritis, but showed better gastrointestinal tolerability.

Ion-pairing high-performance liquid chromatographic method for the detection of N-acetylaspartate and N-acetylglutamate in cerebral tissue extracts.

An ion-pairing high-performance liquid chromatographic method for the determination of N-acetylaspartate and N-acetylglutamate using a C-18 column and a UV detection at 210 nm wavelength, by means of a diode array detector, is presented. A buffer containing 2.8 mM tetrabutylammonium hydroxide, 25 mM KH(2)PO(4), 1.25% methanol, pH 7. 00, is utilized for the isocratic separation of these N-acetylated amino acids, at a flow rate of 1 ml/min and a column temperature of 23 degrees C. The suitability of this chromatographic separation (without additional chromatographic steps prior to HPLC assay) to monitor variations both of N-acetylaspartate and of N-acetylglutamate in perchloric acid brain extracts from rats subjected to the impact acceleration model of diffuse brain injury is also reported. According to the data presented, this HPLC method allows the separation of the two N-acetylated amino acids considered from the many possible interfering compounds, commonly present in extracts of cerebral tissue, which have high extinction coefficients at 210 nm wavelength. Values of N-acetylaspartate and N-acetylglutamate determined by this method showed that cerebral trauma negatively affects both compounds, according to the severity of trauma itself.

Clinical and gastroscopic evaluation of amtolmetin guacyl versus diclofenac in patients with rheumatoid arthritis.

AIM: Amtolmetin guacyl (2-[2[1-methyl-5-(4-methylbenzoyl) pyrrol-2-yl] acetamido] acetic acid 2-methoxyphenyl ester) is a recently developed drug which, in preliminary studies, has shown effective anti-inflammatory properties with improved gastrointestinal safety. Our study was designed to investigate the efficacy and tolerability of amtolmetin guacyl 600 mg bid when compared to diclofenac 50 mg tid for 4 weeks. PATIENTS AND METHODS: A total of 64 patients aged 18-80 years, suffering from rheumatoid arthritis for more than 6 months and American Rheumatism Association functional class I, II or III were randomized in a double blind manner to amtolmetin guacyl or diclofenac for 4 weeks. Clinical and endoscopic evaluation were performed at baseline and at the end of the treatment. The mucosa was graded by means of a rating system emphasizing mucosal erosions. Only patients with endoscopy grade 0-1 entered the trial. RESULTS: The median post-treatment endoscopy injury scores were 0 (range 0-4) in the amtolmetin guacyl-treated patients and 2 (range 0-4) in the diclofenac-treated patients (p = 0.005). There were nine gastric ulcers: 1/32 (3%) in the amtolmetin guacyl group and 8/32 (25%) in the diclofenac group (p < 0.05; 95% confidence interval, -30-5%). 16/32 (50%) patients in amtolmetin guacyl group and 8/32 (25%) in diclofenac group had normal gastroduodenal findings (score = 0) (p < 0.05; 95% confidence interval, 5-50%). In patients with a history of peptic ulcer, a recurrence of gastric damage (score 3-4) was observed in 18% in the amtolmetin guacyl and in 53% in the diclofenac group (p < 0.05). The incidence of gastrointestinal symptoms did not differ in the two groups. Amtolmetin guacyl significantly reduced the number of swollen and painful joints, and the functional disability index; diclofenac significantly reduced the number of painful joints and the functional disability index score (p = ns). CONCLUSIONS: Amtolmetin guacyl effectively controlled the symptoms of rheumatoid arthritis, with very limited gastric toxicity. If these findings are confirmed on a wider scale, the drug might become a valid alternative to current therapies, especially for patients at risk, such as those with rheumatoid arthritis simultaneously requiring steroids and second-line drugs, or those with a history of peptic ulcer.

Peptic ulcer therapy with cimetidine versus tripotassium dicitrato bismuthate in rheumatoid arthritis patients undergoing chronic NSAID treatment.

AIM: To compare the efficacy of cimetidine and tripotassium dicitrato bismuthate (TDB) in arthritic patients who had developed gastric (GU) or duodenal (DU) ulceration while taking non-steroidal anti-inflammatory drugs (NSAIDs). METHODS: Eighty-six rheumatoid arthritis (RA) patients affected by endoscopically proven DU (n = 44) or GU (n = 42), and on chronic NSAID therapy which was not suspended during anti-ulcer therapy, were randomized to cimetidine (400 mg t.d.s.) or TDB (120 mg q.d.s.). A repeat endoscopy was planned after 4 weeks (and 8 weeks, in case of failed healing). The patients who were unhealed after 8 weeks of therapy were allocated to the alternative anti-ulcer drug for a further 8 weeks without interrupting the anti-inflammatory therapy. RESULTS: At week 4 of therapy. 14/24 (58%) DU and 9/20 (45%) GU patients treated with cimetidine were healed, compared with 12/20 (60%) and 10/22 (45%) TDB-treated patients (N.S.). At week 8 of therapy, the DU healing rates were 15/24 (63%) with cimetidine and 14/20 (70%) for TDB. The corresponding GU healing rates were 12/20 (60%) with cimetidine and 13/22 (60%) for TDB (N.S.). At week 16, complete healing with cimetidine was observed in 67% of DU and 57% of GU patients unhealed with TDB; the corresponding figures in the patients crossed to TDB were 83% for DU and 63% for GU patients (N.S. vs. cimetidine). CONCLUSIONS: No statistically significant difference was found between the healing activities of cimetidine and TDB in rheumatoid arthritis patients with peptic ulcer who did not interrupt their NSAID treatment for arthritis. This trial showed that the continued consumption of NSAIDs appears to slow the ulcer healing process, especially in GU patients.

Prevention of gastroduodenal damage with omeprazole in patients receiving continuous NSAIDs treatment. A double blind placebo controlled study.

AIM: The aim of this study was to compare omeprazole (20 mg once daily) with placebo in the long-term prevention of gastroduodenal lesions induced by indomethacin, diclofenac and ketoprofen. PATIENTS AND METHODS: 114 patients with arthritic disorders and requiring indomethacin, diclofenac or ketoprofen were randomized in a double blind manner to receive omeprazole-20 mg once daily- or identical placebo for three weeks. The gastroduodenal mucosa damage was scored according to a 0-4 point endoscopic scale. RESULTS: Of the 114 patients, 103 (50 in the omeprazole group, 53 in the placebo group) were submitted to endoscopy, while 11 patients dropped out for non-medical reasons. At the final endoscopy, 26/57 (46%) of omeprazole group, and 20/57 (35%) of the placebo group had normal gastroduodenal mucosa (score = 0) (p ns; 95% IC -0.073 + 0.284). A gastric ulcer was observed in 7/57 (12%) patients, all in the placebo group (p < 0.01 vs omeprazole); 2 patients (1 in the omeprazole group and 1 in the placebo group) developed a duodenal ulcer. Dyspeptic symptoms developed in 10% of the patients treated with omeprazole and 29% of those receiving placebo (p ns). CONCLUSIONS: Omeprazole, 20 mg once daily, provides effective prophylactic therapy in patients at risk of developing NSAID-associated gastric and duodenal ulcer.