3D Multicellular Tumor Spheroids in a Microfluidic Droplet System for Investigation of Drug Resistance.

A three-dimensional (3D) tumor spheroid model plays a critical role in mimicking tumor microenvironments in vivo. However, the conventional culture methods lack the ability to manipulate the 3D tumor spheroids in a homogeneous manner. To address this limitation, we developed a microfluidic-based droplet system for drug screening applications. We used a tree-shaped gradient generator to control the cell density and encapsulate the cells within uniform-sized droplets to generate a 3D gradient-sized tumor spheroid. Using this microfluidic-based droplet system, we demonstrated the high-throughput generation of uniform 3D tumor spheroids containing various cellular ratios for the analysis of the anti-cancer drug cytotoxicity. Consequently, this microfluidic-based gradient droplet generator could be a potentially powerful tool for anti-cancer drug screening applications.

Anti-Arthritic and Analgesic Effect of NDI10218, a Standardized Extract of Terminalia chebula, on Arthritis and Pain Model.

The fruit of Terminalia chebula Retzius has been used as a panacea in India and Southeast Asia but its biological activities have not been fully elucidated. Here we report anti-arthritic and analgesic effect of NDI10218, a standardized ethanol extract of Terminalia chebula, on collagen-induced arthritis and acetic acid-induced writhing model, respectively. Arthritis was induced in DBA/1J mice by immunizing bovine type II collagen and mice were treated with NDI10218 daily for 5 weeks after the onset of the disease. NDI10218 reduced the arthritis index and blocked the synovial hyperplasia in a dose-dependent manner. The serum levels of pro-inflammatory cytokines TNF-α, IL-6, and IL-1ß were significantly reduced in mice treated with NDI10218. Production of the inflammatory IL-17, but not immunosuppressive IL-10, was also inhibited in splenocytes isolated from NDI10218-treated arthritis mice. Administration of NDI10218 markedly decreased the number of T cell subpopulations in the regional lymph nodes of the arthritis mice. Finally, NDI10218 reduced the number of abdominal contractions in acetic acid-induced writhing model, suggesting an analgesic effect of this extract. Taken together, these results suggest that NDI10218 can be a new therapeutic candidate for the treatment of rheuma-toid arthritis.

Usefulness of the NMP22BladderChek Test for Screening and Follow-up of Bladder Cancer.

PURPOSE: We evaluated the usefulness of the nuclear matrix protein 22 BladderChek (NMP22BC) test for the screening and follow-up of bladder cancer. MATERIALS AND METHODS: From February 2006 to September 2009, we enrolled 1,070 patients who had hematuria or who were being followed up for bladder cancer. We compared the sensitivity and specificity of the NMP22BC test with those of urine cytology. RESULTS: The sensitivity of the NMP22BC test (77.5%) was significantly higher than that of urine cytology (46.3%). The specificity of the NMP22BC test was 88.8%, compared with 97.9% for urine cytology. The sensitivity of the NMP22BC test (81.8%) in non-muscle-invasive bladder cancer was higher than that of cytology (36.4%). However, the sensitivity of the NMP22BC test and of urine cytology in invasive bladder cancer were 57.1% and 92.9%, respectively. The sensitivity of the NMP22BC test was higher for low-grade bladder cancer (83.9%) than for high-grade (62.5%), and the sensitivity of cytology was higher for high-grade bladder cancer (66.7%) than for low-grade (37.5%). Follow-up bladder cancer was detected in 262 patients. The sensitivity of the NMP22BC test in that group (72.7%) was decreased and the specificity (91.7%) was increased. The sensitivity of cytology (54.5%) in the follow-up group was increased and the specificity (95.6%) was decreased. The presence of pyuria was significantly associated with the lower specificity of the NMP22BC test. CONCLUSIONS: The greater sensitivity of the NMP22BC test may be more useful for the diagnosis of non-muscle-invasive bladder cancer and low-grade bladder cancer than for the diagnosis of invasive or high-grade bladder cancer. If the NMP22BC test is performed in the absence of pyuria, it may play a compensatory role for urine cytology.

Screening and isolation of a natural dopamine D1 receptor antagonist using cell-based assays.

To develop a cell-based assay to screen for human dopamine D(1) receptor agonists or antagonists from medicinal plant extracts, a stable Chinese hamster ovary (CHO) cell line (CHO-D1R) expressing the human dopamine D(1) receptor was established using an expression vector containing a scaffold attachment region (SAR) element. CHO-D1R cells showed specific binding to [(3)H]-SCH23390 with high affinity (K(d)=1.47+/-0.17 nM) and dose-dependent responses for the dopamine-mediated stimulation of cAMP concentrations (EC(50)=20.6+/-1.44 nM). The screening of medicinal plant extracts using cell-based cAMP assays revealed that an extract of Gleditsia sinensis Lam., which is known to be rich in saponin, had strong antagonist activity for the D(1) receptor. From the activity-guided fractionation and chemical structural analysis of the G. sinensis extract, a compound called gleditsioside F was isolated and was identified to have antagonist activity for the D(1) receptor. Gleditsioside F showed very effective D(1) antagonist activity by inhibiting ligand binding to the D(1) receptor as well as by inhibiting dopamine-mediated increases in cAMP concentration.

Shengmai-san-mediated enhancement of regenerative responses of spinal cord axons after injury in rats.

Shengmai-san (SMS) is a traditional Chinese medicine used to treat diverse symptoms including cardiovascular and neurological disorders. Here we investigated the effects of SMS on regenerative responses of spinal cord axons in rats that were given contusion injury at the lower thoracic level. The injury cavity was confined to a restricted area by SMS treatment, and the signals of glial scar protein chondroitin sulphate proteoglycan (CSPG) and inflammatory cell marker protein CD11beta were heavily observed within the injury cavity in SMS-treated animals. Anterograde tracing of DiI-labeled corticospinal tract (CST) axons revealed increases in collateral arborization around and within the injury cavity and caudal elongation by SMS treatment. Furthermore, SMS treatment facilitated neurite elongation of dorsal root ganglion (DRG) sensory neurons that were co-cultured with non-neuronal cells prepared from injured spinal cord. Phospho-Erk1/2 was strongly induced in both spinal cord and motor cortical areas after spinal cord injury (SCI), and it was further unregulated in the motor cortex by SMS treatment. In contrast, upregulation of cell division cycle 2 (Cdc2) production by SMS treatment was limited to a local, SCI area. These data suggest that SMS may play an active role in regenerative responses and facilitate axonal regrowth after SCI.

A case of facial myofascial pain syndrome presenting as trigeminal neuralgia.

Facial pain has many causes, including idiopathic factors, trigeminal neuralgia, dental problems, temporomandibular joint disorders, cranial abnormalities, and infections. However, the clinical diagnosis of facial pain is sometimes difficult to establish because clinical manifestations commonly overlap. The diagnosis of trigeminal neuralgia is based solely on clinical findings. Therefore, a careful evaluation of the patient history and a thorough physical examination are essential. This case describes a patient with facial myofascial pain syndrome involving the right zygomaticus, orbicularis oculi, and levator labii muscles, which presented as trigeminal neuralgia.

Effect of muscle relaxant on entropy during propofol anesthesia.

BACKGROUND: The purpose of this study was to investigate whether muscle relaxant affect the values of Entropy, response entropy (RE) or state entropy (SE) during propofol anesthesia. METHODS: Eighty patients (ASA I) scheduled for elective surgery under general anesthesia were randomly assigned to four groups. Anesthesia was maintained at a SE value of 80 (80 +/- 2) using target controlled infusion (TCI) of propofol. After maintaining SE 80 for 5 min, vecuronium 0.1 mg/kg was injected intravenously in group I and same volume of normal saline was intravenously injected in group II. After maintaining SE 60 for 5 min, vecuronium 0.1 mg/kg was injected intravenously in group III and same volume of normal saline was injected intravenously in group IV. The mean arterial pressure, heart rate, SE and RE were measured before anesthetic induction and up to 5 min after vecuronium or normal saline injection in each group. RESULTS: SE and RE were not changed in group II, but significantly decreased in group I (P < 0.05, respectively). In group III and IV, SE and RE were not changed in both groups. There were no significant hemodynamic changes among the four groups. CONCLUSIONS: These results suggest that the effect of muscle relaxant on Entropy vary according to the baseline values of RE or SE during propofol anesthesia.

Tracheal laceration detected by high end-tidal CO2 during endoscopic thyroidectomy.

Endoscopic thyroidectomy is frequently used for cosmetic reasons, such as reducing cervical scarring. Subcutaneous gas insufflation with CO2 is needed to maintain the surgical space, and optimal surgical techniques and careful attention are required when conducting this procedure due to the limited space available for the endoscopic instruments. We report here a case of a tracheal laceration with a tear in the cuff of a reinforced tube, which was detected by an abrupt increase in end-tidal CO2 to 90 mmHg. Reintubation was achieved using a tube exchanger and the patient was effectively ventilated without complications.

Immune-suppressive activity of punicalagin via inhibition of NFAT activation.

Since T cell activation is central to the development of autoimmune diseases, we screened a natural product library comprising 1400 samples of medicinal herbal extracts, to identify compounds that suppress T cell activity. Punicalagin (PCG) isolated from the fruit of Punica granatum was identified as a potent immune suppressant, based on its inhibitory action on the activation of the nuclear factor of activated T cells (NFAT). PCG downregulated the mRNA and soluble protein expression of interleukin-2 from anti-CD3/anti-CD28-stimulated murine splenic CD4+ T cells and suppressed mixed leukocytes reaction (MLR) without exhibiting cytotoxicity to the cells. In vivo, the PCG treatment inhibited phorbol 12-myristate 13-acetate (PMA)-induced chronic ear edema in mice and decreased CD3+ T cell infiltration of the inflamed tissue. These results suggest that PCG could be a potential candidate for the therapeutics of various immune pathologies.

Suppression of the onset and progression of collagen-induced arthritis by chebulagic acid screened from a natural product library.

OBJECTIVE: Chebulagic acid (CHE) from the immature seeds of Terminalia chebula was identified from a natural product library as a potent suppressor of T cell activity. This study examined the effectiveness of CHE against the onset and progression of collagen-induced arthritis (CIA) in mice. METHODS: Arthritis was induced in DBA/1J mice by subcutaneous immunization with bovine type II collagen on days 0 and 21. CHE was administered intraperitoneally for 3 weeks, either as prophylaxis (10 or 20 mg/kg) before disease onset or as therapy (20 mg/kg) after disease onset. Clinical scores, serum antibody levels, and cytokines were measured, and flow cytometric analysis and real-time reverse transcription-polymerase chain reaction were performed to evaluate the knee joints of mice with CIA. RESULTS: In both the prophylactic and therapeutic CHE dosing models, all clinical scores, serum levels of total and anticollagen IgG, and levels of interleukin-10 (IL-10) and IL-6 were reduced, while serum levels of transforming growth factor beta (TGFbeta) were markedly elevated. The number of granulocytes was reduced, but the proportion of CD4+,CD25+ T cells was greater in the knee joints of CHE-treated CIA mice. Expression of Foxp3 and TGFbeta messenger RNA was also augmented significantly in the knee joints of CHE-treated CIA mice in the therapeutic dosing model. CONCLUSION: CHE significantly suppressed the onset and progression of CIA in mice. Immune suppression via the induction of TGFbeta and CD4+,CD25+ T cells may represent a new strategy in the development of therapies for managing rheumatoid arthritis and other inflammatory diseases.

Decrease in genu of the corpus callosum in medication-naïve, early-onset dysthymia and depressive personality disorder.

BACKGROUND: The purpose of this study was to detect differences in regional areas of the corpus callosum (CC) in subjects with early-onset minor depression (dysthymia or depressive personality disorder) and healthy comparison subjects. Based on previous reports that have suggested reduced frontal lobe volume and reduced hemispheric lateralization in the frontal regions of the brain in depression, we hypothesized that the area of the CC that interconnects the frontal regions of the brain, i.e., the genu, will be smaller compared to that of healthy comparison subjects. METHODS: Forty female subjects with early-onset dysthymia or depressive personality disorder, as defined by the Structured Clinical Interview for DSM-III-R and the Diagnostic Interview for Depressive Personality, respectively, and age- and gender-matched healthy comparison subjects (n = 42) were recruited (age: 21.4 +/- 2.1 and 20.9 +/- 2.8 years, respectively). All subjects were psychotropic medications-naïve and right-handed. A 1.5T GE Sigma scanner was used to acquire 124 1.5-mm-thick contiguous coronal images. Midsagittal slice images were carefully selected from reconstructed magnetic resonance images both from native and stereotaxic space to measure seven regional areas of the CC. RESULTS: There were significant diagnosis by CC region interactions [F(6,480) = 4.06, p <.001; F(6,480) = 3.30, p =.003, native and stereotaxic space, respectively]. Early-onset minor depression subjects had a 9.9% (native space) and 6.9% (stereotaxic space) smaller genu of the CC compared to the healthy comparison subjects (the Newman-Keuls post hoc test, p =.005 and.019, native and stereotaxic space, respectively). Early-onset minor depression subjects also had a 7.8% smaller posterior midbody relative to the comparison subjects (the Newman-Keuls post hoc test, p =.033) only in the native space. Severity of current depressive symptoms or duration of illness did not correlate with the size of the genu or the posterior midbody parts of the CC. CONCLUSIONS: These results suggest frontal lobe structural, and possibly functional, abnormalities in the brain in young female adults with a milder spectrum of depression, i.e., DSM-IV early-onset dysthymia or depressive personality disorder. The present findings point out the possible role of frontal lobe abnormality in pathophysiology of early-onset minor depression.