Determining the pharmacologic window of bisphosphonates that mitigates severe injury-induced osteoporosis and muscle calcification, while preserving fracture repair.

Following severe injury, biomineralization is disrupted and limited therapeutic options exist to correct these pathologic changes. This study utilized a clinically relevant murine model of polytrauma including a severe injury with concomitant musculoskeletal injuries to identify when bisphosphonate administration can prevent the paradoxical decrease of biomineralization in bone and increased biomineralization in soft tissues, yet not interfere with musculoskeletal repair. INTRODUCTION: Systemic and intrinsic mechanisms in bone and soft tissues help promote biomineralization to the skeleton, while preventing it in soft tissues. However, severe injury can disrupt this homeostatic biomineralization tropism, leading to adverse patient outcomes due to a paradoxical decrease of biomineralization in bone and increased biomineralization in soft tissues. There remains a need for therapeutics that restore the natural tropism of biomineralization in severely injured patients. Bisphosphonates can elicit potent effects on biomineralization, though with variable impact on musculoskeletal repair. Thus, a critical clinical question remains as to the optimal time to initiate bisphosphonate therapy in patients following a polytrauma, in which bone and muscle are injured in combination with a severe injury, such as a burn. METHODS: To test the hypothesis that the dichotomous effects of bisphosphonates are dependent upon the time of administration relative to the ongoing biomineralization in reparative bone and soft tissues, this study utilized murine models of isolated injury or polytrauma with a severe injury, in conjunction with sensitive, longitudinal measure of musculoskeletal repair. RESULTS: This study demonstrated that if administered at the time of injury, bisphosphonates prevented severe injury-induced bone loss and soft tissue calcification, but did not interfere with bone repair or remodeling. However, if administered between 7 and 21 days post-injury, bisphosphonates temporally and spatially localized to sites of active biomineralization, leading to impaired fracture callus remodeling and permanence of soft tissue calcification. CONCLUSION: There is a specific pharmacologic window following polytrauma that bisphosphonates can prevent the consequences of dysregulated biomineralization, yet not impair musculoskeletal regeneration.

Clinical outcome of endoscopic mucosal resection for esophageal squamous cell cancer invading muscularis mucosa and submucosal layer.

When a tumor invades the muscularis mucosa and submucosal layer (T1a-MM and T1b in Japan), esophageal squamous cell cancer poses 10-50% risk of lymph node metastasis. By this stage of esophageal cancer, surgery, although very invasive, is the standard radical therapy for the patients. Endoscopic mucosal resection (EMR) is the absolutely curable treatment for cancer in the superficial mucosal layer. Because of its minimal invasiveness, the indications of EMR may be expanded to include the treatment of T1a-MM and T1b esophageal carcinoma. To date, the clinical outcomes of EMR for T1a-MM and T1b patients have not been fully elucidated. Here, the retrospective analysis of the clinical outcomes is reported. Between January 1994 and December 2007, 247 patients underwent EMR at Kanagawa Cancer Center. Of these individuals, 44 patients with 44 lesions fulfilled the following criteria: (i) extended EMR treatment for clinical T1a-MM and T1b tumor; (ii) diagnosis of clinical N0M0; and (iii) follow up for at least 1 year, and negative vertical margin. These patients were reviewed for their clinical features and outcomes. Statistical analyses were performed by the Kaplan-Meier methods, the Chi-square test, and the Cox proportional hazard model. P-value of <0.05 was considered statistically significant. The data were analyzed in February 2009. Based on the informed consent and their general health conditions, 44 patients decided the following treatments immediately after the EMR: 2 underwent surgery, 1 underwent adjuvant chemotherapy, and 41 selected follow up without any additional therapy. Of the 41 patients, 20 selected this course by choice, 12 because of severe concurrent diseases, 2 because of poor performance status, and 7 because of other multiple primary cancers. Twelve patients died; two were cause specific (4.5%), eight from multiple primary cancers, one from severe concurrent diseases, and one from unknown causes. No critical complications were noted. Median follow-up time was 51 months (12-126). Five patients ultimately developed lymph node metastasis. One patient with adjuvant chemotherapy required surgery, and another was treated with chemotherapy whose subsequent death was cause specific. The other three patients received chemoradiotherapy and have not shown cause-specific death. Overall and cause-specific survival rates at 5 years were 67.3% and 91.8%, respectively. Among 41 patients treated by EMR alone, only one died from primary esophageal cancer (2.4%), and overall and cause-specific survival rates at 5 years were 75.6% and 97.6%, respectively. Multivariate analysis revealed that severe concurrent diseases including multiple primary cancers and the administration of 5-fluorouracil-based chemotherapy for multiple primary cancers significantly influenced survival (P= 0.025, hazard ratio [HR] 13.1 [95% confidence interval 1.5-114]) and (P= 0.037, HR 0.213 [95% confidence interval 0.05-0.914]), respectively. Eight and six patients developed metachronous esophageal squamous cell cancer and local recurrence, respectively. With the exception of one patient, they could be retreated endoscopically. EMR is a reasonable option for the patients with T1a-MM and T1b esophageal carcinoma without clinical metastasis, especially for the individuals with severe concurrent diseases. The prognostic factors for the benefit of EMR in such cases should be further examined.

Effectiveness of the computed analysis of electroglottographic signals in muscle tension dysphonia.

OBJECTIVE: To investigate the usefulness of electroglottography (EGG) parameters in the diagnosis and estimation of efficacy of voice therapy for muscle tension dysphonia (MTD). PATIENTS AND METHODS: Nineteen MTD participants, an equivalent number of dysphonic ('organic') patients with vocal fold lesions and as many normal speakers were enrolled. Acoustic (Ac) and EGG signals during sustained phonation were recorded simultaneously. The period and amplitude perturbation quotient of both signals, the closed quotient (CQ) of EGG signals (mean CQ) and its standard deviation (CQSD) were calculated, and subsequently compared among the three groups. These parameters in the MTD group were compared before and after voice therapy. RESULTS: The perturbation measures of both signals in the MTD group were either as high as or significantly higher than those in the organic group or the control group, respectively. Both the Ac and EGG parameters after therapy significantly decreased. The CQSD, but not mean CQ, also decreased after therapy. CONCLUSION: EGG parameters related to the regularity of vocal fold vibration, but not to the degree of vocal fold contact (mean CQ), are useful for the diagnosis and estimation of voice therapy outcome in MTD.

New wrinkles on wrinkling: an 8-year longitudinal study on the progression of expression lines into persistent wrinkles.

BACKGROUND: While cumulative lifetime sun exposure is well recognized as having an important role in the progression of facial wrinkling, the role of facial expression has largely been overlooked, in part due to the lack of comprehensive longitudinal data on the change in both expression lines and persistent wrinkles with age. OBJECTIVES: To track the detailed pattern of facial wrinkling in the same group of people over several years and to verify that expression lines evolve into persistent wrinkles. In addition, to identify factors predictive of a faster or slower rate of wrinkling. METHODS: Standardized images were captured at baseline and at 8 years of 122 women (ages 10-72 years, skin types I-VI) with and without a smiling expression. The wrinkle pattern with expression at baseline was compared with the pattern without expression at 8 years. Severity of facial wrinkling was quantified using computer-based image analysis. Skin colour, hydration, sebum and pH were measured at baseline. A structured questionnaire captured demographic and lifestyle data at baseline and at 8 years. RESULTS: Each subject's unique pattern of persistent facial wrinkling observed without expression at year 8 was predicted by the pattern of lines observed with a smiling expression at baseline. Having a drier, more alkaline stratum corneum, a lighter complexion, being middle-aged (40s) or becoming menopausal were associated with faster persistent wrinkling. CONCLUSIONS: Repeated skin flexure during facial expression causes persistent wrinkles. The pattern of expression lines predicts the pattern of future persistent wrinkles. Certain intrinsic and extrinsic factors are not causative, but influence the rate, of facial wrinkling.

Metabolic tumor volume of metastatic lymph nodes and survival after total laryngectomy in laryngeal and hypopharyngeal cancer.

OBJECTIVES: The prognostic value of metabolic tumor volume (MTV) in locally advanced laryngeal or hypopharyngeal cancer is established in the setting of chemoradiotherapy, while it remains unknown in the setting of upfront total laryngectomy. MATERIALS AND METHODS: We retrospectively analyzed 88 patients receiving total laryngectomy and neck dissection, using Cox regression models. RESULTS AND CONCLUSION: Variables related to metastatic lymph node were associated with overall survival, whereas those related to primary tumor were not. In multivariable models, MTV of metastatic lymph nodes (N-MTV) as a continuous variable (Akaike's information criterion (AIC), 277.5) was equivalent to pathological nodal status (AIC, 278.2; P = 0.40), and superior to pathological nodal classification as an ordinal variable (AIC, 281.4; P < 0.05) in ability of predicting death. The risk of death was increased by 1.2-fold (95% confidence interval (CI), 1.0-1.4; P = 0.03) every 10-ml increment of N-MTV, while patients with pN+ disease were at a higher risk of death by 2.9-fold (95% CI, 1.0-12.2; P < 0.05) compared with patients with pN0 disease. Using recursive partitioning analysis (RPA), we classified the patients as having a low, intermediate, or high risk of death on the basis of N-MTV and extranodal extension (ENE). This RPA classification system exhibited greater concordance with overall survival than the classification considering pathological nodal status and ENE (AIC, 275.8 versus 281.4; P = 0.02). In the setting of upfront total laryngectomy, N-MTV is a critical predictor of mortality. A staging system in which N-MTV is incorporated may better inform adjuvant treatment decisions.

The single-prolonged stress paradigm alters both the morphology and stress response of magnocellular vasopressin neurons.

Vasopressin (AVP) plays an important role in anxiety-related and social behaviors. Single-prolonged stress (SPS) has been established as an animal acute severe stress model and has been shown to induce a lower adrenocorticotropic hormone (ACTH) response upon cortisol challenge. Here, we show results from immunoassays for AVP, ACTH, and corticosterone (CORT), and in situ hybridizations for AVP mRNA performed 7 days after SPS exposure. Immunofluorescence for AVP was also performed during the 7-day period following SPS exposure and after an additional forced swimming stress paradigm. We observed that the plasma concentrations of AVP, ACTH, and CORT were not altered by SPS; ACTH content in the pituitary and AVP mRNA expression in the supraoptic nucleus (SON) were significantly reduced by SPS. During the 7-day period following SPS, the intensity of immunoreactivity, the size of the soma, and the immunoreactive optical density of the dendrites of AVP neurons in the SON all increased. An apparent reduction in the intensity of AVP immunoreactivity was observed in the SON at 4 h after additional stress. Additional forced swimming led to a rapid increase in the dendritic AVP content only in the controls and not in the SPS-treated rats. These findings suggest that AVP is a potential biomarker for past exposure to severe stress and that alterations in AVP may affect the development of pathogenesis in stress-related disorders.

Thermodynamics of binding between saccharides and Helix pomatia A hemagglutinin.

Binding constants for the interactions between Helix pomatia A hemagglutinin and the following saccharides were estimated at pH 7.0 and 25 degree C, using the circular dichroism method: N-acetyl-D-galactosamine, 5700 M-1; N-acetyl-D-glucosamine, 1000 M-1; melibiose, 86 M-1; raffinose, 350 M-1. The binding of N-acetyl-D-galactosamine to Helix pomatia A hemagglutinin was investigated in detail, using the circular dichroism and fluorescence methods, over the temperature range 5-50 degree C and pH range 7.0-2.5. The thermodynamic parameters, delta H degree (kcal . mol-1), delta G degree (kcal . mol-1), and delta S degree (e.u.), for this binding reaction at 25 degrees C were estimated as follows: -11.3, -5.24, -20.3 at pH 7.0; -9.1, -5.71, -11.2 at pH 4.5; -38.3, -3.19, -118 at pH 2.5. The negative values of delta H degrees and delta S degrees at pH 2.5 were especially large. This may be related to the restoration of the Helix pomatia A hemagglutinin molecule or its binding site from an unstable configuration caused by lowering the pH of the reaction medium to the original configuration of the presence of N-acetyl-D-galactosamine.

Expression of cytoplasmic galectin-3 as a prognostic marker in tongue carcinoma.

Galectin-3 is a member of the beta-galactoside-binding mammalian lectin family with affinity to ABH group epitopes, cell surface and extracellular polylactosamine glycans. It has been shown to be involved in differentiation, morphogenesis, tumor progression, and metastasis. Here we questioned the possible involvement of galectin-3 in the neoplastic progression of the tongue epithelium and evaluated its prognostic value in tongue cancer patients. Galectin-3 expression was analyzed by the immunohistochemical method in 77 tongue specimens (54 squamous cell carcinomas and 23 specimens of distinct normal mucosa). Levels of nuclear expression of galectin-3 markedly decreased during the progression from normal to cancerous states (P < 0.0001), while cytoplasmic expression increased (P < 0.0001). Enhanced expression of galectin-3 in the cytoplasm was associated with a reduced disease-free survival of tongue cancer patients. Multivariate analysis identified enhanced expression of cytoplasmic galectin-3 as an independent predictor of disease recurrence (P = 0.0120). These results suggest that the observed translocation of galectin-3 from the nucleus to the cytoplasm during neoplastic progression may serve as a prognostic factor for tongue cancer patients.

[Fungal endocarditis found at onset of lower limb acute aortic occlusion; report of a case].

Fungal endocarditis caused by Candida species is associated with high morbidity and mortality. A combination of surgical resection and antifungal drug therapy is the golden standard for treatment. We reported a case of fungal endocarditis due to Candida lusitaniae found at onset of lower limb acute aortic occlusion cured by emergency operation. This case suggests that Candida endocariditis can be managed medically with antifungal drug therapy in life time.

Galectin-3 maintains the transformed phenotype of thyroid papillary carcinoma cells.

Galectin-3, a beta-galactoside-binding protein, is highly expressed in thyroid papillary carcinomas, while functional relevance of galectin-3 overexpression to the malignant phenotype remains elusive. In the present study we transfected galectin-3 antisense cDNA into the human thyroid papillary carcinoma cell line NPA which expresses an innately high level of galectin-3, and examined the effect of antisense inhibition of galectin-3 expression on the transformed phenotype. There was no difference in anchorage-dependent growth between the antisense clones and either the control or parental clones. In contrast, anchorage-independent growth and saturation density of the antisense clones were significantly suppressed compared to those of either the control or parental clones. These results demonstrate that overexpression of galectin-3 in thyroid papillary carcinoma cells is necessary for the maintenance of transformed phenotype, and suggest galectin-3 as a potential target for therapeutic interventions in the future.

Effect of p-chlorophenylalanine on diazepam withdrawal signs in rats.

The effect of p-chlorophenylalanine (PCPA), a specific serotonin (5-HT) depleter, on diazepam withdrawal signs was studied. Rats were made dependent on diazepam by the chronic administration of this drug in the diet. At the time of diazepam withdrawal, the animals were treated with PCPA (200 mg/kg, IP) or the corresponding vehicle (control). After diazepam withdrawal, the maximal body weight losses of control and PCPA-treated animals were 4.1% and 9.0%, respectively. In naive animals, PCPA did not cause any change in body weight. These results suggest that depletion of central 5-HT by PCPA may potentiate the severity of diazepam withdrawal signs.

Muscimol induces state-dependent learning in Morris water maze task in rats.

Effects of muscimol on the place learning in Morris water maze task were investigated in rats. Rats were given 4 training trials per day with the submerged platform at a fixed location in the maze for 4 days. On day 4, rats were required to swim in the pool without the platform after 4 training trials (probe test). Compared to the saline-treated rats, the rats treated with muscimol on day 1-4 showed no modifications of place learning in the training trials and the probe test. However, in the rats treated with muscimol on day 1-3 and treated with saline on day 4, there was increased latency to reach the platform and reduced duration in the quadrant where the platform had been located on day 4. The increased latency in the training trials and reduced duration in the probe test on day 4 was blocked by bicuculline, when bicuculline and muscimol were co-administered on day 1-3, and saline was injected on day 4. Moreover, in the rats treated with muscimol on day 1-3, co-administration of bicuculline and muscimol on day 4 blocked place learning: increased latency in the training trials and reduced duration in the probe test was observed. These results suggest that muscimol induces state-dependent learning (SDL) in Morris water maze task, and that muscimol-induced SDL is mediated by GABAA receptors.

Involvement of cholinergic systems in the deficit of place learning in Morris water maze task induced by baclofen in rats.

Effects of oxotremorine on the deficit of place learning in the Morris water maze task induced by baclofen and scopolamine were examined to determine the involvement of brain cholinergic systems in the deficit of learning induced by baclofen. Rats were given 4 training trials per day with the submerged platform at a fixed location in the maze for 4 days. On day 4, rats were required to swim in the pool without the platform after the 4th training trial (probe test). Baclofen as well as scopolamine dose-dependently increased the escape latency in the training trials. In the probe test, baclofen as well as scopolamine dose-dependently reduced the duration in the quadrant where the platform had been originally located. Increased latency in the training trials and reduced duration in the probe test induced by scopolamine were dose-dependently attenuated by oxotremorine. Increased latency and reduced duration in the baclofen-treated rats were improved by oxotremorine as well as 2-hydroxysaclofen. Baclofen but not scopolamine induced motor incoordination in the rotarod test. Oxotremorine failed to improve motor incoordination induced by baclofen. These results suggest that cholinergic systems may be involved in the deficit of place learning induced by baclofen, and that the ameliorative effects of oxotremorine may not be due to improvement of motor incoordination.

Involvement of GABA(B) receptor systems in action of antidepressants: baclofen but not bicuculline attenuates the effects of antidepressants on the forced swim test in rats.

Involvement of GABAergic systems in action of antidepressants was examined in the forced swim test in rats. Rats were forced to swim in a cylinder for 15 min on day 1 and for 5 min on day 2. Desipramine, mianserin and buspirone, administered after the 15-min swim session on day 1 and before the 5-min swim test on day 2, dose-dependently decreased the duration of immobility in the swim test on day 2. Baclofen attenuated the decreased duration of immobility induced by desipramine, mianserin and buspirone in the swim test, although baclofen did not affect the duration of immobility when it was injected alone. Muscimol dose-dependently decreased the duration of immobility in the swim test on day 2. Bicuculline antagonized the decreased duration of immobility induced by muscimol. However, bicuculline failed to antagonize the decreased duration of immobility induced by desipramine, mianserin and buspirone. These results suggest that GABA(B) but not GABA(A) receptor systems may be involved in action of antidepressants.

Activation of ventral tegmental GABAB receptors inhibits morphine-induced place preference in rats.

The effect of microinjection of a GABAB receptor agonist, baclofen, into the ventral tegmental area on the rewarding effect of morphine was investigated using the conditioned place preference paradigm in rats. Morphine (1-8 mg/kg, s.c.) caused a dose-related place preference for the drug-associated place. In contrast, microinjection of baclofen (0.1-1 nmol/side) into the ventral tegmental area did not produce a significant preference for either compartment of the test box. Pretreatment with baclofen (0.1-1 nmol/side) into the ventral tegmental area dose dependently suppressed the morphine (8 mg/kg, s.c)-induced place preference. This suppression of the morphine (8 mg/kg, s.c.)-induced place preference by baclofen (1 nmol/side), but not with the GABAA receptor antagonist bicuculline (1 nmol/side). The present results suggest that a decrease in GABAB neurotransmission in the ventral tegmental area, which may be produced via inhibition of a tonic GABAergic input by morphine, may be involved in the expression of the rewarding effect of morphine.

Induction of physical dependence on pentobarbital by new infusion method in the rat.

1. A new intermittent intravenous infusion method was developed for the induction of tolerance to and physical dependence on pentobarbital in rats. 2. Female and male rats were injected with pentobarbital (20 mg/kg/injection) through an implanted intravenous cannula. 3. The rats were allowed to receive an injection after the completion of a fixed amount of behavioral activity counted from the preceding injection, and therefore, the sedative and hypnotic effects of pentobarbital were used as an index in the determination of the injection intervals. 4. During pentobarbital treatment, the number of pentobarbital injections per day rapidly increased and stabilized (approximately 40: male, approximately 30: female) on the third to fifth day. 5. Upon withdrawal, the female and male rats who were maintained on pentobarbital administration of more than 30 and 40 injections/day for approximately 10 days manifested withdrawal signs which included spontaneous convulsion. 6. These results suggest that a new infusion method exists to produce a high degree of physical dependence in rats on a short-acting barbiturates, pentobarbital.

Development of physical dependence on and tolerance to morphine in rats treated with morphine-admixed food.

1. The development process of physical dependence on and tolerance to morphine has been explored in rats treated with morphine-admixed food (0.5 mg/g of food) during 1 to 7 days. 2. In the morphine-treated animals, body weight loss was observed after the abrupt morphine withdrawal. 3. Intensity and time course of the weight loss were correlated to the morphine treatment. 4. On the other hand, the morphine-treated rats showed abnormal behaviors, such as diarrhea, ptosis, teeth chattering, salivation, body shakes, vocalization, nose bleed, irritability, aggression, lacrimation and writhing upon naloxone injection. 5. Loss of body weight, measured 3 hours after naloxone injection, was also correlated to the duration of morphine treatment. 6. Tolerance to the analgesic effect of morphine developed within one day in rats treated with morphine-admixed food. 7. The drug-admixed food ingestion method has the advantage of rapidly inducing a high degree of physical dependence and tolerance without causing morbidity or lethality in animals. It also eliminates the need for excessive handling of animals.

Cytotoxic xanthones from Garcinia hanburyi.

Eleven novel cytotoxic xanthones, gambogin, morellin dimethyl acetal, isomoreollin B, moreollic acid, gambogenic acid, gambogenin, isogambogenin, desoxygambogenin, gambogenin dimethyl acetal, gambogellic acid and hanburin were isolated together with four known xanthones, gambogic acid, isomorellin, morellic acid and desoxymorellin, from the dry latex of Garcinia hanburyi. The structures were elucidated by a detailed spectroscopic analysis.

Antiviral activity of lignans and their glycosides from Justicia procumbens.

Ten antiviral lignans, seven known (justicidins A, B, C and D, diphyllin, diphyllin apioside and diphyllin apioside-5-acetate) and three new compounds, justicidinosides A (justicidin C 6'-O-glucoside), B (justicidin A 6'-O-glucoside) and C (justicidin B 6'-O-glucoside), were isolated from a methanolic extract of the aerial parts of Justicia procumbens var. leucantha. Justicidins A and B, diphyllin, diphyllin apioside and diphyllin apioside-5-acetate showed strong antiviral activity (the MIC were less than 0.25 microgram ml-1, respectively) against vesicular stomatitis virus and low cytotoxicity (the MTC were larger than 31 micrograms ml-1, respectively) against cultured rabbit lung cells (RL-33).

Effect of opioid agonist-antagonist interaction on morphine dependence in rats.

Morphine dependence was induced by treatment with morphine-admixed food (0.25mg/g of food) for 7 days. Withdrawal was precipitated by injecting naloxone (0.5mg/kg, s.c.). Rats treated with morphine exhibited body weight loss upon the naloxone injection. When morphine-dependent rats were injected subcutaneously with morphine, codeine, meperidine and pentazocine 30 min before the naloxone injection, these drugs significantly suppressed the naloxone-precipitated loss of body weight in a dose-dependent manner. However, body weight loss induced through coadministration of naloxone and Mr-2266 BS were not suppressed by morphine pretreatment. These results suggest that opioids protect against naloxone-precipitated loss of body weight, and that mu and kappa opiate receptors play an important role in the protection against naloxone-precipitated withdrawal.