Blue Dye Staining Research Articles

99mTc-dextran lymphoscintigraphy can detect sentinel lymph node in breast cancer patients.

The aim of this study was to retrospectively determine the accuracy and feasibility of using 99mTc-dextran (DX) lymphoscintigraphy for the localization of sentinel lymph nodes (SLNs) in breast cancer patients. The relevant factors affecting lymphoscintigraphy were also investigated. In this study, 235 breast cancer patients underwent 99mTc-DX lymphoscintigraphic imaging and examination by a γ-probe method in combination with blue dye staining to detect SLNs. The detection results were considered in combination with rapid frozen pathology results to determine whether SLN metastasis was positive or negative. SLNs were identified in 191 patients by γ-probe detection among the 202 patients that tested positive by lymphoscintigraphic imaging, a coincidence rate of 94.6%. This suggested that lymph node metastasis had occurred and could be detected using lymphoscintigraphy. The axillary status of the breast cancer patients was also predicted using lymphoscintigraphy and the false-negative rate, sensitivity, specificity and positive predictive value were 13.3% (4/30), 90.7% (39/43), 23.4% (45/192) and 13.5% (21/155), respectively. The age of the patient, menstrual status, tumor location, tumor size, pathological type, preoperative biopsy and neoadjuvant chemotherapy were unrelated to the success of lymphoscintigraphy (P>0.05). 99mTc-DX lymphoscintigraphy is able to exactly determine the location of SLN in breast cancer patients, and can be used for guiding γ-probe methods and sentinel lymph node biopsy.

Phototoxic effect of na-pheophorbide a toward osteosarcoma cells in vitro using a laser diode.

The purpose of this study was to investigate the effectiveness of photodynamic therapy (PDT) with Na-pheophorbide A in anticancer treatment, using osteosarcoma cells in vitro. It has been reported that PDT with chlorophyll derivatives inhibits the proliferation of various cancer cells. However, there have been no reports that have evaluated the effectiveness of PDT in suppressing osteosarcoma cells. Uptake of Na-pheophorbide A into Hu09 cells (osteosarcoma cells) was assayed using fluorescence microscopy following incubation of the cells with 28 μmol/L of Na-pheophorbide A. The viability of Hu09 cells after PDT treatment was assessed using trypan blue dye staining and MTS assays. PDT-induced apoptosis was determined by evaluation of the activity of selected members of the caspase family and by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining of cells. Na-pheophorbide A uptake by cells was rapid, being observed after 60 min of treatment, and Na-pheophorbide A persisted in cells for >24 h. PDT treatment decreased cell viability compared with the control group, indicating high cytocidal activity of PDT. This cytocidal effect was dependent upon drug concentration, light dose, and the number of irradiation times. An increase in the number of cells positive for TUNEL staining and increases in the activity of caspases-3, -8 and -9 were observed in the first 2 h after PDT treatment. A cytotoxic effect of PDT with Na-pheophorbide A on an osteosarcoma cell line in vitro was shown. Caspase activity assays suggested that PDT with Na-pheophorbide A induced an apoptotic change in HuO9 cells, mainly via activation of mitochondrial caspase -9 and -3 pathways.

Localization of sentinel lymph node in breast cancer. A prospective study

IntroductionSentinel Lymph Node Biopsy (SLNB) is the standard of care for staging axillary lymph nodes in women with breast cancer and clinically negative nodes. It is associated with reduced arm morbidity, moderated or severe lymphoedema, and a better quality of life in comparison with standard axillary treatment. Unfortunately, skip metastases makes all minimally invasive approaches, such as axillary sampling, unreliable. The aim of the present clinical prospective study is to evaluate the position of SLN in an important number of cases and establish the real incidence of skip metastases in clinically node-negative patients. Patients and methodsA cohort of 898 female patients with breast carcinoma was considered, from 2001 to 2008. Once SLN was localized, by means of radio-colloid or blue dye staining, and isolated, a biopsy was performed. Only those positive for metastases were submitted to axillary dissection. ResultsOnly in nine cases a SLN was not isolated. We had 819 cases of first level SLN (group A) and 69 cases of second level SLN (group B). Considering all of 889 cases, SLN was localized in the second level in 69 patients (7.8%); but if we consider metastatic SLN alone (340 cases), it was in the second level in 23 subjects (6.8%). In total, we had a positive second level SLN in 2.3% of cases (23/889). ConclusionSecond level SLN could be considered only an anomalous lymphatic axillary drainage and it does not linked to particular histological variants of the primitive tumour. In our study, skip metastases were recognized in only 2.6% of cases, therefore, whenever a SLN is not isolated for any reason, the first level sampling represent a viable operative choice.

Sleep deprivation impairs functional muscle recovery following injury

Introduction Skeletal muscles possess the ability to completely regenerate following muscular injury. Sleep is believed to play an important role in this regenerative process; however, the nature of this role has not been previously tested. Our aim was to investigate the effects of sleep deprivation on molecular, histological and functional indices of muscle repair following myotoxic injury. Materials and methods Male rats were injected with 1.5% bupivacaine into the masseter muscle to induce myotoxic damage. Subjects were either sleep deprived for 8 h during the light period using a forced locomotion activity wheel, or served as activity controls. They were subsequently sacrificed at 2, 7 or 14 days post-injection. Western Blot analysis was used to assay for protein expression of positive (MyoD, myogenin) and negative (myostatin) molecular repair markers. Evans Blue Dye staining for damaged muscle fibres was used to examine histopathology. Functional muscle repair was evaluated after 2 weeks using in situ contractility testing to measure the force–frequency relationship during isometric contractions. Results Sleep deprivation suppressed MyoD protein levels in the masseter at 2 and 7 days, and myogenin at 2 days post-injection, compared to activity controls. Myostatin levels were unaffected. Histopathology revealed no effect of sleep deprivation on the extent of muscle fibre degeneration following injury. The force-frequency curve tended to shift downward and to the right in response to sleep loss, indicating compromised contractile force at moderate to high stimulation frequencies. Conclusion We demonstrate that sleep loss impairs functional recovery of the masseter muscle following myotoxic injury. Specifically, 8 h of sleep deprivation acutely downregulated molecular markers of muscle repair and resulted in contractile function deficits during recovery. Together, these findings suggest that sleep normally plays a permissive role in the regeneration of damaged muscle tissue. Acknowledgements PS thanks the National Sciences and Engineering Research Council of Canada (NSERC) for his Ph.D. funding. This research is supported by funds from Canadian Institutes of Health Research (CIHR) and NSERC .

Abstract 9434: Post-infarct Treatment With Microrna145 Protects the Heart Against Myocardial Ischemia-reperfusion Injury Through Acceleration of Myocyte Autophagy

Background: We previously reported that microRNA145 (miRNA145) attenuates the ischemia-reperfusion injury. However, the mechanism how miRNA145 reduces the myocardial infarct size is still unclear. Objective: We aimed to investigate whether post-infarct treatment with miRNA145 reduces myocardial infarct size and its precise mechanism in a rabbit model of myocardial infarction. Methods: Male Japanese white rabbits underwent 30 min of coronary occlusion followed by 14 days of reperfusion. Rabbits received intravenous injection of saline (control group, n=7) or 0.035 mg/kg of miRNA145 encapsulated with liposome (miRNA145 group, n=7) immediately after reperfusion. At 14 days after reperefusion, rabbits were sacrificed and the hearts were removed. The area at risk and infarct areas were measured by evans blue dye and TTC staining, respectively. The infarct size was calculated as a percentage of the risk area of the left ventricle. Heart tissues were sampled and the morphological changes were investigated by electron microscopy, and the expression of LC3B, an indicator of autophagy, was assessed by western blot analysis. In another series of experiments, rat myoblast H9C2 were treated with 20 nM of miRNA145 and investigated the morphological changes by electron microscopy and the expression of LC3B by western blot analysis. Results: Post-infarct treatment with miRNA145 significantly reduced the myocardial infarct size (16.0±3.1 %) as compared to the control (28.7±6.1 %) group. Post-infarct treatment with miRNA145 significantly improved LV remodeling and improved ±dp/dt as compared to the control group. Both in vitro myoblast and in vivo experiments, treatment with miRNA145 induced autophagic changes such as autophagosome and autolysosome, and activated LC3B. Conclusions: It is suggested that post-infarct treatment with miRNA145 attenuated ischemia-reperfusion injury through acceleration of myocyte autophagy. The miRNA145 may be a promising strategy for treatment of acute myocardial infarction.

Hot or not? The 10% rule in sentinel lymph node biopsy for malignant melanoma revisited

The surgeon needs a practical rule to follow when deciding whether to excise a lymph node during sentinel node biopsy (SLNB). The "10% rule" dictates that all nodes with a radiation count of greater than 10% of the hottest node and all blue nodes should be removed, and this study observes the effects of following this rule in SLNB in melanoma. We reviewed the records of 665 patients with primary melanoma who underwent sentinel lymph node over a 5-year period (2007-2011). 2064 nodes were identified in 898 nodal basins in 665 patients. 141 (21%) patients had at least one positive sentinel node. 105 positive nodal basins were identified in which more than one sentinel node was removed. In 18 of these, a less radioactive node was positive for tumour when the most radioactive node was negative. Of 175 positive nodes 157 (90%) contained blue dye staining. For cases in which the positive sentinel node was not the hottest node, the positive node had apparent blue dye staining in all 18 cases (100%), and was the second hottest node in the basin. In this series removing just the hottest node and all blue nodes would not have missed a single positive basin and would have resulted in a 38% reduction in the number of nodes removed compared to those taken following the 10% rule, without changing the staging in any patient.

Post-infarct treatment with microRNA145 reduces myocardial infarct size through acceleration of myocyte autophagy in rabbits

Background: We previously reported that microRNA145 (miRNA145) attenuates the ischemia-reperfusion injury. However, it is not still unclear how miRNA145 attenuates the ischemia-reperfusion injury. Objective: We aimed to investigate whether post-infarct treatment with miRNA145 reduces myocardial infarct size and its precise mechanism in a rabbit model of myocardial infarction. Methods: Male Japanese white rabbits underwent 30 min of coronary occlusion followed by 14 days of reperfusion. Rabbits received intravenous injection of saline (control group, n=7) or 0.035 mg/kg of miRNA145 encapsulated with liposome (miRNA145 group, n=7) immediately after reperfusion. At 14 days after reperfusion, rabbits were sacrificed and the hearts were removed. The area at risk and infarct areas were measured by evans blue dye and TTC staining, respectively, and the infarct size was calculated as a percentage of the risk area of the left ventricle. In another series of experiments, rat myoblast H9C2 were treated with 20 nM of miRNA145 and investigated the morphological changes by electron microscopy and signals by western blot analysis such as LC3B. Results: Post-infarct treatment with miRNA145 significantly reduced the myocardial infarct size (16.0±3.1%) as compared to the control (28.7±6.1%) group. Post-infarct treatment with miRNA145 significantly improved LV remodeling and improved ±dp/dt as compared to the control group. In the myoblast experiment, treatment with miRNA145 caused autophagic changes such as autophagosome and autolysosome, and caused activation of LC3B. Conclusions: It is suggested that post-infarct treatment with miRNA145 attenuated ischemia-reperfusion injury through acceleration of myocyte autophagy. The miRNA145 may be a promising strategy for treatment of acute myocardial infarction.

Endoplasmic reticulum stress contributes to heart protection induced by cyclophilin D inhibition

Preventing cyclophilin D (cypD) translocation to the inner mitochondrial membrane can limit lethal reperfusion injury through the inhibition of the opening of the mitochondrial permeability transition pore. Inhibition or loss of function of cypD may also result into an endoplasmic reticulum (ER) stress that has been shown to alter cell survival. We therefore questioned whether ER stress might play a role in the protection induced by CypD deficiency or inhibition. CypD-KO and NIM811 (a CypD inhibitor)-treated mice were subjected to a prolonged ischemia-reperfusion (I/R). Area at risk and infarct size was measured using blue dye and triphenyltetrazolium chloride staining. ER stress markers were measured in the hearts during the reperfusion phase. As expected, cypD-KO mice exhibited a decreased infarct size when compared to wild-type mice (8 ± 1 vs. 20 ± 4% of left ventricular weight; p < 0.01). CypD-deficient mice displayed an increased expression of ER stress proteins such as eukaryotic initiation factor 2α (eIF2α) or glucose regulated protein 78 (Grp78 or Bip). The ER stress inhibitor TUDCA prevented the infarct size reduction afforded by the loss of cypD function (mean infarct size averaged 21 ± 4% of LV weight, p < 0.01 vs. cypD-KO). Similar results were obtained when NIM811, an analog of cyclosporine A, was used to pharmacologically (instead of genetically) inhibit cypD function. This study suggests that the ER stress induced by the inhibition of cypD function plays a key role in protecting the heart against lethal ischemia-reperfusion injury.

Human IL10 Gene Repression by Rev-erbα Ameliorates Mycobacterium tuberculosis Clearance

Nuclear receptors modulate macrophage effector functions, which are imperative for clearance or survival of mycobacterial infection. The adopted orphan nuclear receptor Rev-erbα is a constitutive transcriptional repressor as it lacks AF2 domain and was earlier shown to be present in macrophages. In the present study, we highlight the differences in the relative subcellular localization of Rev-erbα in monocytes and macrophages. The nuclear localization of Rev-erbα in macrophages is subsequent to monocyte differentiation. Expression analysis of Rev-erbα elucidated it to be considerably more expressed in M1 phenotype in comparison with M2. Rev-erbα overexpression augments antimycobacterial properties of macrophage by keeping IL10 in a basal repressed state. Further, promoter analysis revealed that IL10 promoter harbors a Rev-erbα binding site exclusive to humans and higher order primates and not mouse, demonstrating a species barrier in its functionality. This direct gene repression is mediated by recruitment of co-repressors NCoR and HDAC3. In addition, our data elucidate that its overexpression reduced the survival of intracellular pathogen Mycobacterium tuberculosis by enhancing phagosome lysosome maturation, an event resulting from IL10 repression. Thus, these findings suggest that Rev-erbα bestows protection against mycobacterial infection by direct gene repression of IL10 and thus provide a novel target in modulating macrophage microbicidal properties.

Ex vivo sentinel node mapping in colon cancer combining blue dye staining and fluorescence imaging

BackgroundThe sentinel lymph node procedure has been proposed to improve nodal staging in colon cancer patients. The aim of this study was to assess the added value of near-infrared (NIR) fluorescence imaging to conventional blue dye staining for ex vivo sentinel lymph node mapping. Materials and methodsWe included 22 consecutive patients undergoing surgery for colon cancer. After tumor resection, we submucosally injected a premixed cocktail of the near-infrared lymphatic tracer HSA800 and blue dye around the tumor for detection of sentinel lymph nodes. We used the Mini-FLARE imaging system for fluorescence imaging. ResultsIn 95% of patients, we identified at least one sentinel lymph node. Overall, a total of 77 sentinel lymph nodes were identified, 77 of which were fluorescent (100%) and 70 of which were blue (91%). Sentinel lymph nodes that were located deeper in the mesenteric fat could easily be located by NIR fluorescence. In four of five patients with lymph node metastases, tumor cells were present in at least one of the sentinel lymph nodes. ConclusionsThis study shows the successful use and added value of the NIR fluorescence tracer HSA800 to conventional blue dye for the ex vivo sentinel lymph node procedure in colon cancer.

Cardiac-specific Coxsackievirus and Adenovirus Receptor (CAR) Deletion Inhibit Enterovirus Infection in Murine Heart

The structure of coxsackievirus and adenovirus receptors CAR is similar to adhesion molecules. In the adult heart, the majority of CAR localizes at the intercalated disc. Germ line CAR deletion induces embryonic lethality at E11.5 with evidence of a cardiac abnormality. The CAR role as a viral receptor is well known; however, its precise function in the heart for enterovirus infection is not clear. To understand the role of CAR in the cardiac myocyte, we generated cardiac-specific CAR knockout mice using a CAR floxed allele and alpha-MHC-Mer CRE Mer mice. Western blot analysis and immunofluorescent stain of ventricles at 6 weeks after 2 weeks tamoxifen administration, CAR expression was significantly decreased in CAR f/f MCM mice but not in CAR f/f mice heart. Enterovirus was intraperitoneally infected into CAR f/f MCM and CAR f/f mice (n=10 each). CAR disruption was dramatically reduced virus infection and replication in the heart but not different in liver, spleen, and pancreas. Cardiac myocyte damage was significantly reduced in the CAR f/f MCM mutant mice by evans blue dye stain. In addition, the CAR f/f MCM mutant mice heart inflammation and fibrosis were decreased in H&E and trichrome stain compare to CAR f/f control mice. CAR expression was required for normal ventricular function, but it is the cause of enterovirus infection. In the adult mice heart, CAR deletion was significantly reduced viral infection, proliferation, and myocarditis. These results suggested that CAR deletion could be useful therapeutic strategy to prevent viral myocarditis.

CALPAIN ACTIVATION CONTRIBUTES TO ISCHAEMIA/REPERFUSION-INDUCED MYOCARDIAL APOPTOSIS

ObjectivesCalpains belong to a family of calcium-dependent intracellular cysteine proteases. Previous studies demonstrated that calpain activation contributes substantially to ischaemia/reperfusion (I/R) injury. However, it remains unclear whether administration of calpain...

Bronchoscopic Observation of Endobronchial Tumor Cells

A 68-year-old woman presented with cough and dyspnea. Chest x-ray film and computed tomography revealed atelectasis in the right upper lobe. She had a history of a bladder neoplasm, which was treated by cystectomy and chemotherapy. Flexible bronchoscopy revealed a polypoid tumor in the right upper lobe bronchus, protruding into the right main bronchus. Further, we observed the surface of the tumor by using the endocytoscopy system. The endocytoscopy system showed oval-shaped tumor cells with prominent nuclei on the surface of the tumor with a methylene blue dye staining. Biopsied specimen of the tumor demonstrated foci of metastatic urothelial carcinoma. These cancer cells were similar to the cell images of endocytoscopy system. This technology may have the potential to provide pathologic diagnosis during bronchoscopy.

Pacing Postconditioning: Impact of Pacing Algorithm, Gender, and Diabetes on Its Myocardial Protective Effects

Pacing postconditioning (PPC) induces cardioprotection. The aim of this study was to determine the optimal pacing algorithm and possible influence of gender and diabetes on PPC. Unprotected regional ischemia for 30min served as negative control and classical PPC (ten cycles of 30s left ventricular pacing alternated with 30s right atrial pacing) as positive control. Area at risk and infarct size were determined by blue dye and triphenyltetrazolium chloride staining. For achieving protection, the minimal number of PPC cycles was seven and the minimal duration of a PPC protocol was 200s. The protective effect of PPC was comparable in male and female hearts, but no protection could be induced by PPC in diabetic hearts. PPC can provide myocardial protection when using at least seven cycles of ventricular pacing. PPC protection is independent of gender, but sensitive to experimental diabetes.

MG53 reduces cell death associated with hemorrhagic shock ischemia/reperfusion (I/R) injury by increasing membrane repair capacity

Physical trauma is a leading cause of death in the United States. While interventions to restore blood pressure and volume following acute trauma are essential, the abrupt restoration of blood flow following ischemia results in significant cell damage and death. As a result, injury caused by a hemorrhagic shock ischemia/reperfusion (I/R) event can have severe consequences, including organ failure and patient death. MG53 is a TRIM family protein which is an essential component of the cell membrane repair machinery in cardiac and skeletal muscle. Following various types of membrane damage, including injury associated with I/R, our research group has shown that MG53 facilitates formation of a membrane repair patch to restore the integrity of the disrupted membrane. Creatine kinase (CK) and lactate dehydrogenase (LDH) assays revealed significantly increased membrane resealing capacity in cultured mammalian cells or isolated mg53−/− flexor digitorum brevis (FDB) muscle fibers in the presence of rhMG53. Histological examination of tissue isolated from mice pretreated with rhMG53 and subjected to hemorrhagic shock reveal a significant decrease in Evan's blue dye staining in multiple tissues, suggesting reduction in necrotic cell death due to enhanced membrane repair. Our results show that recombinant human MG53 (rhMG53) can serve a protective role and mitigate cellular death and damage brought on by I/R following hemorrhagic shock. Research was funded by the Foundation for UMDNJ.

Vitreo-papillary adhesion as a prognostic factor in pseudo- and lamellar macular holes

To determine the incidence of vitreopapillary adhesion (VPA) and to investigate its value as a prognostic factor in the surgical outcome of pseudo- (PMH) and lamellar macular holes (LMH). A total of 76 consecutive patients, diagnosed with PMH (41 eyes) or LMH (35 eyes) were included. Eyes with VPA were alternatively assigned to the surgical or control group. Surgery consisted of a 25G vitrectomy and internal limiting membrane peeling with blue dye staining. There were six visits: baseline, the day of surgery, and 1, 3, 6, and 9 post-operative months. Main outcome measures were the incidence of VPA and changes in the outer retinal layers and visual acuity. VPA was found in 27% (11/41) of patients with PMH and 37% (13/35) with LMH (P=0.03). In presence of VPA, the best-corrected visual acuity (BCVA) improved in the surgery group from 32 ± 8 to 47 ± 8 letters, whereas the control group went from 34 ± 7 to 31 ± 8 letters. The difference in letters between the surgery and control groups was statistically significant for both distance (P=0.032) and near (P=0.04) vision. Intra-retinal cysts were significantly correlated with a poor functional prognosis (P=0.01). We found the presence of focal damage to the outer retinal layers in LMH and PMH. VPA is more frequent in the presence of LMH vsPMH. It significantly influences the tangential forces at the vitreoretinal interface, exacerbating anatomical changes, and worsening the functional prognosis. Functional difference between the surgical and control groups was statistically significant for distant and near vision in presence of VPA.

Membrane Sealant Poloxamer P188 Protects Against Isoproterenol Induced Cardiomyopathy in Dystrophin Deficient Mice

BackgroundCardiomyopathy in Duchenne muscular dystrophy (DMD) is an increasing cause of death in patients. The absence of dystrophin leads to loss of membrane integrity, cell death and fibrosis in cardiac muscle. Treatment of cardiomyocyte membrane instability could help prevent cardiomyopathy.MethodsThree month old female mdx mice were exposed to the β1 receptor agonist isoproterenol subcutaneously and treated with the non-ionic tri-block copolymer Poloxamer P188 (P188) (460 mg/kg/dose i.p. daily). Cardiac function was assessed using high frequency echocardiography. Tissue was evaluated with Evans Blue Dye (EBD) and picrosirius red staining.ResultsBL10 control mice tolerated 30 mg/kg/day of isoproterenol for 4 weeks while death occurred in mdx mice at 30, 15, 10, 5 and 1 mg/kg/day within 24 hours. Mdx mice tolerated a low dose of 0.5 mg/kg/day. Isoproterenol exposed mdx mice showed significantly increased heart rates (p < 0.02) and cardiac fibrosis (p < 0.01) over 4 weeks compared to unexposed controls. P188 treatment of mdx mice significantly increased heart rate (median 593 vs. 667 bpm; p < 0.001) after 2 weeks and prevented a decrease in cardiac function in isoproterenol exposed mice (Shortening Fraction = 46 ± 6% vs. 35 ± 6%; p = 0.007) after 4 weeks. P188 treated mdx mice did not show significant differences in cardiac fibrosis, but demonstrated significantly increased EBD positive fibers.ConclusionsThis model suggests that chronic intermittent intraperitoneal P188 treatment can prevent isoproterenol induced cardiomyopathy in dystrophin deficient mdx mice.

Sentinel lymph node in colorectal cancer — 5 years follow up

AbstractTo assess the impact of micrometastases in sentinel and non-sentinel lymph nodes on long-term survival rates of patients treated for colorectal cancer (CRC). Data of 57 patients diagnosed with CRC and treated in the Department of Surgical Oncology in Gdansk in the years 2002–2006 were retrospectively analyzed. Clinico-histopathological data were analyzed using chi-square tests. The effect on long-time survival rates was analyzed using Kaplan-Meier survival probability estimates. Identification of the SLN was performed using the blue dye staining method. All regional lymph nodes were subject to standard histopathological examination. Additionally in 32(56.14%) patients whose nodes were found negative for metastases on standard staining further immunohistochemical analyses were performed. In the analyzed group SLNB was performed in 42(73.7%) patients with colon cancer and in 15(26.3%) with rectal cancer. Identification of the SLN was possible in 45(78.9%) patients. The sensitivity of SLNB was 33%. False negatives were found in 66%. SLNB is a feasible method in CRC patients. We presume that lack of micrometastases in the SLN and non-SLN cannot be regarded as a prognostic factor.

Adenosine A3receptor stimulation induces protection of skeletal muscle from eccentric exercise-mediated injury

Effective therapy to reduce skeletal muscle injury associated with severe or eccentric exercise is needed. The purpose of this study was to determine whether adenosine receptor stimulation can mediate protection from eccentric exercise-induced muscle injury. Downhill treadmill exercise (-15 degrees ) was used to induce eccentric exercise-mediated skeletal muscle injury. Experiments were conducted in both normal wild-type (WT) mice and also in beta-sarcoglycan knockout dystrophic mice, animals that show an exaggerated muscle damage with the stress of exercise. In the vehicle-treated WT animals, eccentric exercise increased serum creatine kinase (CK) greater than 3-fold to 358.9 +/- 62.7 U/l (SE). This increase was totally abolished by stimulation of the A(3) receptor. In the dystrophic beta-sarcoglycan-null mice, eccentric exercise caused CK levels to reach 55,124 +/- 5,558 U/l. A(3) receptor stimulation in these animals reduced the CK response by nearly 50%. In the dystrophic mice at rest, 10% of the fibers were found to be damaged, as indicated by Evans blue dye staining. While this percentage was doubled after exercise, A(3) receptor stimulation eliminated this increase. Neither the A(1) receptor agonist 2-chloro-N(6)-cyclopentyladenosine (0.05 mg/kg) nor the A(2A) receptor agonist 2-p-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine (0.07 mg/kg) protected skeletal muscle from eccentric exercise injury in WT or dystrophic mice. The protective effect of adenosine A(3) receptor stimulation was absent in mice, in which genes for phospholipase C beta2/beta3 (PLCbeta2/beta3) and beta-sarcoglycan were deleted. The present study elucidates a new protective role of the A(3) receptor and PLCbeta2/beta3 and points to a potentially effective therapeutic strategy for eccentric exercise-induced skeletal muscle injury.

Tc-99m diphosphonate as a potential radiotracer to detect sentinel lymph nodes in patients with breast cancer.

To evaluate the potential of Tc-99m diphosphonate as a tracer for sentinel lymph node biopsy in breast cancer. Lymphoscintigraphs of 35 patients (50.9 ± 10.2years) with breast cancer were acquired after administering a subareolar intradermal injection of Tc-99m diphosphonate 18h before surgery. Static images were taken within 15min (early phase) and 15h after injection (delayed phase). The lymphoscintigraphic identification rate was defined as the percentage of subjects studied with visible foci at axillae. Sentinel lymph node biopsies were performed using a gamma probe and by blue dye injection. Any node that was radioactive or stained with blue dye was labeled as a sentinel lymph node. Lymph nodes without radioactivity or blue dye staining were defined as non-sentinel lymph nodes. The intraoperative identification rate was defined as the percentage of patients with a radioactive sentinel lymph node. Percentages of lymphoid cells expressing S-100, CD83, and CD1a were compared. The lymphoscintigraphic identification rate was 94.3% (33/35) during the early phase and 96.9% (31/32) during the delayed phase, whereas the intraoperative identification rate was 94.3% (33/35). The mean percentages of lymphoid cells that stained positively for S-100 or CD83 were lower in sentinel lymph nodes than in non-sentinel lymph nodes (1.5% vs. 9.0% for S-100, and 4.5% vs. 9.3% for CD83, respectively, p = 0.0286). The mean percentages of lymphoid cells in sentinel lymph nodes and non- sentinel lymph nodes expressing CD1a were 3.3% and 7.0%, respectively (p = ns). Tc-99m diphosphonate can reliably detect regional lymph nodes in breast cancer.