Profiling of Patients with Temporomandibular Disorders: Experience of One Tertiary Care Center

ObjectivesThe aim of this study was to assess typical and most prevalent characteristics of patients suffering from temporomandibular disorders (TMD) by a retrospective assessment of their medical records.Material and MethodsDemographic data and data on the characteristics of TMD were collected from the existing medical documentation of 304 TMD patients (250 females and 54 males) who had been referred to the Department of Dentistry, Clinical Hospital Center Zagreb from October 2016 to October 2020 due to temporomandibular pain. For the purpose of analysis, three age groups were formed: i) “children and adolescents” (up to 19 years of age); ii) “middle age” (from 20 to 50 years of age); iii) “older age” (>50 year- olds). A two-step cluster analysis was performed with the aim of classifying TMD patients into homogenous groups.ResultsThe mean age of patients whose data were included in the study was 33.8 ± 16.66, with a significantly higher age in the group of women (p<0.001). Most of the patients had chronic pain (67.4%), with the ratio in favor of chronic patients being significantly higher in women than in men (p=0.001). Data on parafunctional behavior were confirmed in 14.5% of patients. Data on the onset of symptoms during/just after orthodontic treatment were present in 14.5% of patients. Data on spontaneous pain, assessed with a visual analogue scale, were recorded in 87 patients, with a mean of 6.14 ± 1.79 and with the highest pain in the “older age” group. Physical therapy was the most common therapeutic modality (56.3%) followed by an occlusal splint (40.5%). The analysis revealed 5 different clusters in the TMD patient data set.ConclusionsOur results are largely in line with current epidemiological knowledge on TMD. Women predominated in all age groups and most of the patients experienced chronic pain. Classifying patients into homogeneous groups using the clustering method could provide better identification of subgroups of conditions that mainly occur together in these patients, thus providing the basis for more specific management.     

Sequestration of a highly reactive intermediate in an evolving pathway for degradation of pentachlorophenol

Microbes in contaminated environments often evolve new metabolic pathways for detoxification or degradation of pollutants. In some cases, intermediates in newly evolved pathways are more toxic than the initial compound. The initial step in the degradation of pentachlorophenol by Sphingobium chlorophenolicum generates a particularly reactive intermediate; tetrachlorobenzoquinone (TCBQ) is a potent alkylating agent that reacts with cellular thiols at a diffusion-controlled rate. TCBQ reductase (PcpD), an FMN- and NADH-dependent reductase, catalyzes the reduction of TCBQ to tetrachlorohydroquinone. In the presence of PcpD, TCBQ formed by pentachlorophenol hydroxylase (PcpB) is sequestered until it is reduced to the less toxic tetrachlorohydroquinone, protecting the bacterium from the toxic effects of TCBQ and maintaining flux through the pathway. The toxicity of TCBQ may have exerted selective pressure to maintain slow turnover of PcpB (0.02 s⁻¹) so that a transient interaction between PcpB and PcpD can occur before TCBQ is released from the active site of PcpB.Anthropogenic compounds used as pesticides, solvents, and explosives often persist in the environment and can cause toxicity to humans and wildlife. In some cases, microbes have evolved the ability to partially or completely degrade such compounds. Complete degradation of anthropogenic compounds requires the evolution of new metabolic pathways that convert these compounds into metabolites that can be used by existing metabolic networks. The sequence of steps in new degradative pathways is dictated by the repertoire of enzymes in exposed microbes and/or microbial communities that can catalyze newly needed reactions. The serendipitous assembly of new degradation pathways can result in the production of intermediates that are toxic due to their intrinsic chemical reactivity or adventitious interactions between these intermediates and cellular macromolecules. The toxicity of such intermediates can pose a significant challenge to the evolution of efficient pathways for the degradation of anthropogenic compounds (1).The pathway for degradation of pentachlorophenol (PCP) by Sphingobium chlorophenolicum (Fig. 1) (2, 3) includes the remarkably toxic intermediate tetrachlorobenzoquinone (TCBQ). The LD₅₀ for Escherichia coli protoplasts is <1 µM (4). TCBQ is the most toxic of a set of 14 quinones (5), with an LD₅₀ of 18 µM for primary rat hepatocytes. (For comparison, the LD₅₀ for benzoquinone is 57 µM and for 2,3,4,6-tetramethylbenzoquinone is 800 µM.) TCBQ is a potent alkylating agent, capable of forming covalent adducts with proteins (6) and with the nucleobases of DNA (7). Thiols such as glutathione react with TCBQ particularly rapidly (3, 6). Depletion of glutathione eliminates a critical cellular mechanism for protection against alkylating agents and oxidative damage. Further, TCBQ can generate hydroxyl radicals by its interaction with hydrogen peroxide (8). Thus, TCBQ is a devastating toxin due to a plethora of molecular mechanisms.Open in a separate windowFig. 1.The initial steps in the pathway for degradation of PCP in S. chlorophenolicum via the intermediates TCBQ and TCHQ are highlighted in green. Hydroxylation of TCP by PcpB forms TCHQ directly.The pathway for degradation of PCP in S. chlorophenolicum is believed to be still in the process of evolving because PCP was introduced as a pesticide less than 100 y ago (9, 10) and some of the enzymes in the pathway function quite poorly (3, 11). The intermediacy of a dangerous compound such as TCBQ in a newly evolving pathway brings up a number of interesting questions. First, why has the pathway evolved via a route that involves TCBQ? Second, how does the bacterium protect itself from the toxic effects of TCBQ? And third, does the intermediacy of TCBQ explain the inefficiency of PCP degradation by S. chlorophenolicum?The answer to the first question emerges from a consideration of the enzymatic repertoire that exists to initiate degradation of naturally occurring phenols. Phenols are common natural products, produced by a range of organisms from microbes to insects and by degradation of lignin. Cleavage of phenols under aerobic conditions requires the introduction of a second hydroxyl group. This reaction can be carried out by cytochrome P450 enzymes or flavin monooxygenases. PCP hydroxylase (PcpB), the first enzyme in the pathway for degradation of PCP, is a flavin monooxygenase (3) and likely originated from an enzyme that hydroxylated a naturally occurring phenol. Hydroxylation of phenols at a position carrying a hydrogen results in formation of a catechol or hydroquinone rather than a benzoquinone (Fig. 1). However, when a good leaving group such as chloride is present at the position of hydroxylation, the unstable intermediate eliminates HCl to form a benzoquinone.The answer to the second question is suggested by the observation that microbes that generate benzoquinones from phenols typically have a reductase that converts a benzoquinone to a hydroquinone. The gene encoding the reductase is located immediately downstream of the gene encoding the hydroxylase (12). Examples are found in the pathways for degradation of 2,4,6-trichlorophenol (TriCP), p-nitrophenol and o-nitrophenol in Cupriavidus necator JMP134 (13), Pseudomonas sp. Strain WBC-3 (14), and Alcaligenes sp. strain NyZ215 (15), respectively. Like these microbes, S. chlorophenolicum contains a gene encoding a reductase (PcpD) immediately downstream of the gene encoding the hydroxylase (PcpB). We have previously shown that pcpD is essential for survival of S. chlorophenolicum at high concentrations of PCP but is not essential for survival at high concentrations of 2,3,5,6-tetrachlorophenol (TCP) (2). In this work, we have addressed the mechanism of that protection. As shown in Fig. 1, hydroxylation of TCP produces tetrachlorohydroquinone (TCHQ) whereas hydroxylation of PCP produces the more toxic TCBQ. PcpD is an efficient TCBQ reductase. However, its ability to prevent the escape of TCBQ to the solvent before it is reduced to TCHQ is an even more critical aspect of its function.Our results provide insights into the answer to the third question. Because PcpB and PcpD do not form a stable complex, the rate of their encounter is limited by diffusion. The slow rate of turnover by PcpB [at 0.02 s⁻¹ (3), it is the rate-limiting step of the pathway (4)] may be a result of selective pressure to prevent production of TCBQ at a rate faster than it can be intercepted by PcpD. Thus, the efficiency of the interaction between PcpB and PcpD may limit the efficiency of the degradation of PCP and prevent selection of a more catalytically robust PcpB.     

Na+ channel modulation and force-frequency relationship in human myocardium

The enhancement of force of contraction (FOC) following increasing frequencies of stimulation is an important mechanism of positive inotropy in human myocardium. The present study aimed to investigate the influence of alterations in Na⁺ influx on FFR in human myocardium. Isometric FOC of electrically stimulated right auricular trabeculae (AUT, n=12) from human nonfailing hearts (n=8) was measured at different stimulation rates (0.5-3 Hz) under control conditions, after increasing Na⁺ influx by the addition of (±)BDF 9148 (BDF, 3 μmol l^-1) and after decreasing Na⁺ influx by the addition of lidocaine (LIDO, 10 μmol l^-1). Additionally, the rate dependent changes in diastolic tension (DT) were measured in all experiments. Under control conditions FOC increased with increasing frequencies of stimulation. The rate at which maximal FOC was observed (SFmax) was 2.0±0.2 Hz and maximal increase in FOC (PIEmax) by increasing frequency of stimulation was +1.5±0.5 mN. After increase of Na⁺ influx by BDF (3 μmol l^-1) SFmax was decreased to 0.8±0.1 Hz (p<0.05 versus control) and PIEmax was +0.1±0.3 mN (p<0.05). When Na⁺ influx was diminished by LIDO (10 μmol l^-1) SFmax and PIEmax were increased compared to control (2.4±0.1 Hz and +4.1±0.9 mN, p<0.05 versus control). The diastolic tension (DT) of AUT at 3 Hz was not changed at higher rates in the control group and after application of LIDO (10 μmol l^-1), whereas after enhancement of Na⁺ influx by BDF there was an increase in DT of +0.7±0.2 at 3Hz (p<0.05 versus control and LIDO). An enhanced Na⁺ influx leads to a decrease in the optimal frequency and to a smaller force potentiation by higher stimulation rates which could be at least partly due to incomplete relaxation at higher frequencies, whereas a reduced Na⁺ influx is followed by opposite alterations. It is concluded that besides Ca²⁺ handling also Na⁺ influx and Na⁺ homeostasis might determine the frequency-induced force potentiation in human myocardium. Thus, the negative FFR in diseased human myocardium might result from changes in cellular Ca²⁺ or Na⁺ regulatory sites. Received: 20 November 1996 / Accepted: 21 February 1997     

Altered brain activation pattern associated with drug-induced attenuation of enhanced depression-like behavior in rats bred for high anxiety.

BACKGROUND: The enhanced depression-like behavior in the forced swim test displayed by rats selectively bred for high anxiety-related behavior (HAB) as compared with their low anxiety counterparts (LAB) is abolished by chronic paroxetine treatment. The aim of the present study was to identify neuronal substrates underlying this treatment response in HABs. METHODS: The HAB rats received paroxetine (10 mg/kg/day) for 24 days via drinking water, and drug-induced modulation of neuronal activation patterns in response to forced swimming was mapped with the expression of the immediate early gene c-Fos as marker. RESULTS: Chronic paroxetine treatment reduced the immobility scores during forced swimming, confirming the previously observed antidepressant-like effect in these animals, and attenuated the forced swim-induced c-Fos response in a restricted set (11 of 70) of brain areas. These included limbic areas such as the prelimbic cortex, parts of the amygdala, the bed nucleus of the stria terminalis, dorsal hippocampus, dorsal lateral septum as well as hypothalamic and hindbrain areas (dorsolateral periaqueductal gray [PAG], locus coeruleus). Untreated LAB rats, which displayed low depression-like behavior comparable to that of treated HABs, also showed low swim stress-induced c-Fos response in most of these same areas, further supporting an association of attenuated neuronal excitability in the identified areas with attenuated depression-like behavior. CONCLUSIONS: These findings indicate that modulation of neuronal activation in a restricted set of defined, mainly limbic as well as selected hypothalamic and hindbrain areas by paroxetine treatment is associated with the reduction of enhanced depression-like behavior in a psychopathological animal model.     

Photodynamic therapy agent with a built-in apoptosis sensor for evaluating its own therapeutic outcome in situ

Identifying the extent of apoptosis in cells or tissues after cancer therapy in real time would be a powerful firsthand tool for assessing therapeutic outcome. We combined therapeutic and imaging functions in one agent, choosing photodynamic therapy (PDT) as an appropriate cancer treatment modality. This agent induces photodamage in irradiated cells and simultaneously identifies apoptotic cells by near-infrared fluorescence. This photodynamic therapy agent with a built-in apoptosis sensor (PDT-BIAS) contains a fluorescent photosensitizer used as an anticancer drug, connected to a fluorescence quencher by a caspase-3 cleavable peptide linker. We demonstrated that cleavage of the peptide linker by caspase-3, one of the executioner caspases involved in apoptosis, results in a detectable increase of fluorescence in solution and in cancer cells after PDT treatment. The apoptosis involvement and drug effectiveness were confirmed by Apoptag and cell viability (MTT) assays supporting the ability of PDT-BIAS to induce and image apoptosis in situ.     

A phase I Study with oral SU5416 in patients with advanced solid tumors: A drug inducing its clearance

Summary Vascular endothelial growth factor (VEGF) is a potent stimulant of angiogenesis. SU5416, is a small molecule tyrosine kinase inhibitor, and a potent inhibitor of VEGF- mediated Flk-1 receptor signaling. Intravenous agent SU5416 has shown evidence of biological activity against a variety of tumor types. The current intravenous dosing regimen is not optimal for long-term administration, which is needed for optimal efficacy. The aim of this study was to evaluate the safety profile and pharmacokinetics of a Nanocrystal Colloidal Dispersion^TM (NCD) SU5416 formulation in humans. Patients with advanced and/or metastatic solid organ tumors were included in the trial; various SU5416 regimens were tested for tolerability, safety and were evaluated concerning pharmacokinetics. The results of this study indicate that induction of clearance after oral dosing of NCD SU5416 in humans occurs and is greater than following IV administration. It has been confirmed that SU5416 is a high clearance compound, also as an oral NCD formulation. The NCD formulation was well tolerated, but no effective drug serum levels could be achieved. These data help to understand the ADME (Absorption, Distribution, Metabolism, Excretion) properties of indoline chemical class compounds. The lessons learned should be applied in the development of next generation indoline anti-angiogenic and anti-tumor compounds.     

Increased incidence of apparently life-threatening events due to supine position

Gastro-oesophageal reflux (GOR) has a high prevalence in infancy. The supine position is among numerous aggravating factors. The exact relationship between GOR and apparently life-threatening events (ALTE) is not clear, although it has been repeatedly investigated. In 1992 the worldwide Back to Sleep campaign was implemented, which had a dramatic effect on the incidence of sudden infant death syndrome (SIDS) with a drop of 50%. Although the vast majority of children now sleep on their back, the effect of this position on ALTE has not been studied. In this retrospective study, we aim to define the potential association between GOR and ALTE. We hypothesise that the incidence of ALTE has increased since the 1992 recommendation. No bias in the population's selection was introduced, as our centre is the only one for paediatric emergencies in the county. A total of 107 children presenting with ALTE were identified during the study period (1987-99). A pH study was performed in the 75 patients presenting with ALTE in the last 6 years of the study (1994-99). Neither morbidity nor mortality was noted in a long-term 4-year follow-up. Our present results show that the frequency of ALTE increased sevenfold (P < 0.005) between 1992 and 1999. The ALTE episodes took place significantly more often in the post-prandial period. The prevalence of GOR was much higher in patients presenting with ALTE (nearly 75%) when compared with the general population. Furthermore, on medical treatment for GOR, very few patients presented with a second episode of ALTE. Consequently it is thought that GOR and ALTE are linked and that ALTE patients would benefit from GOR treatment. The worldwide marked decrease in SIDS since the implementation of the supine position possibly masks the negative effect of an increase in ALTE.     

Notch1 signaling promotes primary melanoma progression by activating mitogen-activated protein kinase/phosphatidylinositol 3-kinase-Akt pathways and up-regulating N-cadherin expression

Cellular signaling mediated by Notch receptors results in coordinated regulation of cell growth, survival, and differentiation. Aberrant Notch activation has been linked to a variety of human neoplasms. Here, we show that Notch1 signaling drives the vertical growth phase (VGP) of primary melanoma toward a more aggressive phenotype. Constitutive activation of Notch1 by ectopic expression of the Notch1 intracellular domain enables VGP primary melanoma cell lines to proliferate in a serum-independent and growth factor-independent manner in vitro and to grow more aggressively with metastatic activity in vivo. Notch1 activation also enhances tumor cell survival when cultured as three-dimensional spheroids. Such effects of Notch signaling are mediated by activation of the mitogen-activated protein kinase (MAPK) and Akt pathways. Both pathways are activated in melanoma cells following Notch1 pathway activation. Inhibition of either the MAPK or the phosphatidylinositol 3-kinase (PI3K)-Akt pathway reverses the Notch1 signaling-induced tumor cell growth. Moreover, the growth-promoting effect of Notch1 depends on mastermind-like 1. We further showed that Notch1 activation increases tumor cell adhesion and up-regulates N-cadherin expression. Our data show regulation of MAPK/PI3K-Akt pathway activities and expression of N-cadherin by the Notch pathway and provide a mechanistic basis for Notch signaling in the promotion of primary melanoma progression.