Usefulness of High Suction Pressure for Sufficient Tissue Collection During Endobronchial Ultrasound Guided Transbronchial Needle Aspiration

Introduction

The optimal suction pressure during endobronchial ultrasound guided transbronchial needle aspiration (EBUS-TBNA) remains to be determined. The aim of this study was to compare suction pressures for performance in collecting sufficient tissue specimens from mediastinal and hilar lymph nodes during EBUS-TBNA.

Methods

Retrospective analysis of consecutive patients with mediastinal and hilar lymphadenopathy who underwent EBUS-TBNA over a 3-year period. Results from patients who underwent EBUS-TBNA using a dedicated 20-mL VacLoc (Merit Medical Systems, Inc, South Jordan, UT) syringe (conventional method, group C) were compared with results from patients in whom a disposable 30-mL syringe (high pressure group, group H) was used. The yield for sufficient histologic specimen retrieval and amount of tissue obtained were compared between the 2 groups.

Results

Of 178 patients who underwent EBUS-TBNA, 131 had lung cancer confirmed by EBUS-TBNA: 35 in group C and 96 in group H. There were 7 patients in group C and 6 in group H who received final diagnoses by cytology alone. There were 28 in group C and 90 in group H who were diagnosed by both cytology and histology. There was a statistically significant difference between the groups in terms of the rate of sufficient sampling for histological specimens (p = 0.04). The H group revealed a tissue area approximately twice that of the C group (p = 0.003). There were no major procedure-related complications in either group.

Conclusion

Higher suction pressures with larger syringe volumes during EBUS-TBNA may be useful for safely collecting sufficient tissue specimens.     

Involvement of TRPM2 in Peripheral Nerve Injury-Induced Infiltration of Peripheral Immune Cells into the Spinal Cord in Mouse Neuropathic Pain Model

Recent evidence suggests that transient receptor potential melastatin 2 (TRPM2) expressed in immune cells plays an important role in immune and inflammatory responses. We recently reported that TRPM2 expressed in macrophages and spinal microglia contributes to the pathogenesis of inflammatory and neuropathic pain aggravating peripheral and central pronociceptive inflammatory responses in mice. To further elucidate the contribution of TRPM2 expressed by peripheral immune cells to neuropathic pain, we examined the development of peripheral nerve injury-induced neuropathic pain and the infiltration of immune cells (particularly macrophages) into the injured nerve and spinal cord by using bone marrow (BM) chimeric mice by crossing wildtype (WT) and TRPM2-knockout (TRPM2-KO) mice. Four types of BM chimeric mice were prepared, in which irradiated WT or TRPM2-KO recipient mice were transplanted with either WT-or TRPM2-KO donor mouse-derived green fluorescence protein-positive (GFP⁺) BM cells (TRPM2^BM+/Rec+, TRPM2^BM–/Rec+, TRPM2^BM+/Rec–, and TRPM2^BM–/Rec– mice). Mechanical allodynia induced by partial sciatic nerve ligation observed in TRPM2^BM+/Rec+ mice was attenuated in TRPM2^BM–/Rec+, TRPM2^BM+/Rec–, and TRPM2^BM–/Rec– mice. The numbers of GFP⁺ BM-derived cells and Iba1/GFP double-positive macrophages in the injured sciatic nerve did not differ among chimeric mice 14 days after the nerve injury. In the spinal cord, the number of GFP⁺ BM-derived cells, particularly GFP/Iba1 double-positive macrophages, was significantly decreased in the three TRPM2-KO chimeric mouse groups compared with TRPM2^BM+/Rec+ mice. However, the numbers of GFP^–/Iba1⁺ resident microglia did not differ among chimeric mice. These results suggest that TRPM2 plays an important role in the infiltration of peripheral immune cells, particularly macrophages, into the spinal cord, rather than the infiltration of peripheral immune cells into the injured nerves and activation of spinal-resident microglia. The spinal infiltration of macrophages mediated by TRPM2 may contribute to the pathogenesis of neuropathic pain.     

Met Is the Most Frequently Amplified Gene in Endometriosis-Associated Ovarian Clear Cell Adenocarcinoma and Correlates with Worsened Prognosis

Clear cell adenocarcinoma of the ovary (OCC) is a chemo-resistant tumor with a relatively poor prognosis and is frequently associated with endometriosis. Although it is assumed that oxidative stress plays some role in the malignant transformation of this tumor, the characteristic molecular events leading to carcinogenesis remain unknown. In this study, an array-based comparative genomic hybridization (CGH) analysis revealed Met gene amplification in 4/13 OCC primary tumors and 2/8 OCC cell lines. Amplification of the AKT2 gene, which is a downstream component of the Met/PI3K signaling pathway, was also observed in 5/21 samples by array-based CGH analysis. In one patient, both the Met and AKT2 genes were amplified. These findings were confirmed using fluorescence in situ hybridization, real-time quantitative PCR, immunoblotting, and immunohistochemistry. In total, 73 OCC cases were evaluated using real-time quantitative PCR; 37.0% demonstrated Met gene amplification (>4 copies), and 8.2% had AKT2 amplification. Furthermore, stage 1 and 2 patients with Met gene amplification had significantly worse survival than patients without Met gene amplification (p<0.05). Met knockdown by shRNA resulted in reduced viability of OCC cells with Met amplification due to increased apoptosis and cellular senescence, suggesting that the Met signaling pathway plays an important role in OCC carcinogenesis. Thus, we believe that targeted inhibition of the Met pathway may be a promising treatment for OCC.     

NT-702 (parogrelil hydrochloride, NM-702), a novel and potent phosphodiesterase inhibitor, improves reduced walking distance and lowered hindlimb plantar surface temperature in a rat experimental intermittent claudication model

NT-702 (parogrelil hydrochloride, NM-702), 4-bromo-6-[3-(4-chlorophenyl)propoxy]-5-[(pyridin-3-ylmethyl)amino]pyridazin-3(2H)-one hydrochloride, a novel phosphodiesterase (PDE) inhibitor synthesized as a potent vasodilatory and antiplatelet agent, is being developed for the treatment of intermittent claudication (IC) in patients with peripheral arterial disease. We assessed the efficacy of NT-702 in an experimental IC model as compared with cilostazol and additionally investigated the pharmacological property in vitro and ex vivo. NT-702 selectively inhibited PDE3 (IC(50)=0.179 and 0.260 nM for PDE3A and 3B) more potently than cilostazol (IC(50)=231 and 237 nM for PDE3A and 3B) among recombinant human PDE1 to PDE6. NT-702 inhibited in vitro human platelet aggregation induced by various agonists (IC(50)=11 to 67 nM) and phenylephrine-induced rat aortic contraction (IC(50)=24 nM). Corresponding results for cilostazol were 4.1 to 17 microM and 1.0 microM, respectively. NT-702 (3 mg/kg or more) significantly inhibited ex vivo rat platelet aggregation after a single oral dose. For cilostazol, 300 mg/kg was effective. In a rat femoral artery ligation model, NT-702 at 5 and 10 mg/kg repeated oral doses twice a day (BID) for 13 days significantly improved the reduced walking distance while the lowered plantar surface temperature was improved at 2.5 mg/kg and more. Cilostazol also improved the walking distance and surface temperature at 300 mg/kg BID but significant difference was only observed for surface temperature on day 8. These results suggest that NT-702 can be expected to have therapeutic advantage for IC.     

Interactions responsible for secondary structure formation during folding of equine beta-lactoglobulin

Equine beta-lactoglobulin forms a compact intermediate at an acidic pH (A state). It also forms an expanded and helical conformation at low temperatures (C state). The structure of a single disulfide mutant C66A/C160A is similar to the A state in the presence of salts, while it is similar to the C state at low anion concentrations. We have investigated the temperature-dependent change in the secondary structure using circular dichroism and proline scanning mutagenesis. At low anion concentrations, the helical content increased linearly as temperature decreased. In the presence of salts, the A state was cooperatively transformed into the C state at low temperatures. This suggests the importance of hydrophobic interactions for stabilizing the A state. Peptides encompassing native-like and non-native alpha-helices were synthesized to investigate the interactions responsible for helix formation in the A and C states. These did not form stable helices, indicating that not only the helices in the A state but also the helices in the C state are stabilized by long-range interactions. A longer fragment, CHIBL, which encompasses the structured region in the A and C states, showed a helical structure. Proline-substituted mutants of CHIBL showed CD spectral changes similar to the corresponding mutants of the full-length protein in the C state. Therefore, CHIBL has a structure similar to the corresponding region of the full-length protein in the C state. This result indicates that interactions responsible for helix formation in the C state reside in the sequence of CHIBL, and that the sequences outside CHIBL are essential for secondary structure formation in the A state.     

Lack of promoting effects of chronic exposure to 1.95-GHz W-CDMA signals for IMT-2000 cellular system on development of N-ethylnitrosourea-induced central nervous system tumors in F344 rats

The present study was performed to evaluate effects of a 2-year exposure to an electromagnetic near-field (EMF) equivalent to that generated by cellular phones on tumor development in the central nervous system (CNS) of rats. For this purpose, pregnant F344 rats were given a single administration of N-ethylnitrosourea (ENU) on gestational day 18. A total of 500 pups were divided into five groups, each composed of 50 males and 50 females: Group 1, untreated controls; Group 2, ENU alone; Groups 3 to 5, ENU + EMF (sham exposure and two exposure levels). A 1.95-GHz wide-band code division multiple access (W-CDMA) signal, which is a feature of the International Mobile Telecommunication 2000 (IMT-2000) cellular system was employed for exposure of the rat head starting from 5 weeks of age, 90 min a day, 5 days a week, for 104 weeks. Brain average specific absorption rates (SARs) were designed to be .67 and 2.0 W/kg for low and high exposures, respectively. The incidence and numbers of brain tumors in female rats exposed to 1.95-GHz W-CDMA signals showed tendencies to increase but without statistical significance. Overall, no significant increase in incidences or numbers, either in the males or females, was detected in the EMF-exposed groups. In addition, no clear changes in tumor types in the brain were evident. Thus, under the present experimental conditions, exposure of heads of rats to 1.95-GHz W-CDMA signals for IMT-2000 for a 2-year period was not demonstrated to accelerate or otherwise affect ENU-initiated brain tumorigenesis.