Apoptosis index correlates with chemotherapy efficacy and predicts the survival of patients with gastric cancer.

The objectives of this study are to investigate the apoptotic changes in gastric adenocarcinoma following neoadjuvant chemotherapy and to illuminate its correlation with response. One 67 gastric cancer patients with cT2-4 or TanyN1-3M0 between January 2006 and December 2007 were included. All patients had previous gastrectomy and D2 lymphadenectomy with curative intent performed. A total of 12 cycles of preoperative mFOLFOX7 chemotherapy was recommended for all patients, and 83 patients received only adjuvant chemotherapy. Resected specimens were subjected to in situ TUNEL assay and scanned with Applied Imaging Ariol SL-50. Apoptosis index (AI) was significantly higher in the patient received preoperative chemotherapy (CS group) than in the patient did not (S group). In the CS group, AI was found to have a strong positive correlation with the pathological response (rho = 0.403, P = 0.0002). A ROC curve presented a score of 49.4 as the AI cutoff value for response, dividing the CS group into two subgroups with significantly different prognoses (P = 0.003) and further allowing identification of 8 patients with significantly better prognosis out of 48 patients evaluated as grades 1a-b according to a pathological response evaluation (P = 0.022). In conclusion, AI is correlated with efficacy of preoperative chemotherapy and prognosis of gastric cancer patients following neoadjuvant chemotherapy. An AI cutoff value for response may be used as a complementary approach to current pathological response evaluations to help identify potential responders.

Apoptosis during the development of pelvic floor muscle in anorectal malformation rats.

PURPOSE: Fecal incontinence and constipation still remain as major postoperative complications after procedures for anorectal malformations (ARM). The striated muscle complex (SMC) is one of the most important factors that influence defecation. Previous studies have demonstrated different degrees of the muscle complex dysplasia dependent on the complexity of ARM. To explore the mechanisms of maldevelopment of SMC in ARM, apoptosis was investigated during pelvic floor muscle development in rat embryos with ARM. METHODS: Anorectal malformations in rat embryos were induced by treating pregnant rats with ethylenethiourea on the 10th embryonic day (E10). Normal and ARM rat embryos from E16 to E21 were serial-sectioned transversely or sagittally, and SMCs were dissected and snap frozen. TdT mediated dUTP Nick Ending Labeling (TUNEL) staining and DNA ladder analysis were performed to identify apoptosis and expression of Bax/Bcl-2 were confirmed with immunohistochemical staining and Reverse Transcription-Polymerase Chain Reaction (RT-PCR) analysis. RESULTS: Hypoplastic and disordered SMC sling shifted cephalad, ventrally, and converged inferior to the rectourethral fistula and infiltrated connective tissue in ARM embryos. In the normal group, TUNEL-positive cells became evident on E17; sporadic positive staining was mainly localized in 2 areas as follows: the junction area between SMC and bulbocarvernosus muscle and posterior to the rectum where bilateral SMC converged. In the ARM group, massive positive staining of nuclei was observed from E16 to E21 and was mainly distributed in the dorsal part of the SMC. Electrophoresis of DNA samples yielded a "ladder" pattern of migration both in normal and the ARM group from E17 to E21, the ladders were stronger in the ARM group. In both groups, the expression of Bax/Bcl-2 was detectable on E17, the immunoreactivity increased on E19 and E21. Compared with the normal group, the expression of Bax was increased, whereas Bcl-2 was declined in the ARM group. Significant upregulation of Bax messenger RNA (mRNA) levels and downregulation of Bcl-2 mRNA levels were observed in ARM embryos. CONCLUSIONS: In the current study, abnormal apoptosis and disturbed expression of Bax/Bcl-2 were identified during SMC development in ARM embryos. It is suggested that precocious, excessive, and dislocated apoptosis might be a fundamental pathogenesis for the maldeveloped SMC in ARM rats. The temporospatial expressions of Bax/Bcl-2 indicate they may have an important role in the regulation of apoptosis of SMC.

The clinical utility of sperm DNA integrity testing.

Sperm DNA damage is more common in infertile men and may contribute to poor reproductive performance. However, current methods for evaluating sperm DNA integrity do not reliably predict treatment outcomes, and no treatment for abnormal DNA integrity has proven clinical value.

DNA end labelling (TUNEL) in a 3 year old girl with Leigh syndrome and prevalent cortical involvement.

Neuropathological study of a 3 1/2 year old girl with familial Leigh syndrome who also harboured a rare ATPase gene mutation disclosed extensive and unusual lesions in the cerebral cortex, despite a typical histological pattern. Early lesions in the periacqueductal grey matter of the brainstem, characterised by capillary congestion and initial regressive neuronal changes, were also observed, along with TUNEL reactive neuronal cells showing morphological signs typical of apoptosis in cortical areas with neuronal cell loss. The finding of lesions in atypical brain areas and for the first time, very early regressive neuronal phenomena, suggest that early changes in crucial brain areas may have been a cause of death. The abundance of TUNEL positive nuclei in cortical areas in the present case suggests that the apoptosis may be involved in the mechanism of neuronal death in Leigh syndrome.

Evaluation of cell proliferation, apoptosis, and angiogenesis in transitional cell carcinoma of the renal pelvis and ureter.

OBJECTIVES: To investigate cell proliferation, apoptosis, and angiogenesis and their roles in transitional cell carcinoma (TCC) of the renal pelvis and ureter. METHODS: Formalin-fixed and paraffin-embedded tissue blocks from 42 patients with TCC of the renal pelvis and ureter were studied. Cell proliferation was assessed by Ki-67 immunostaining, and the proliferation index (PI) was expressed as a percentage of Ki-67-positive cells. Apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), and the apoptotic index (AI) was expressed as a percentage of TUNEL positive cells. Angiogenesis was evaluated by CD31 immunostaining, and microvessel density (MVD) was expressed as the average of the microvessel count. RESULTS: The PI ranged from 5.9% to 48.0% (median 20.03%), AI from 1.0% to 4.2% (median 2.26%), and MVD from 16.0 to 146.0 (median 56.88) in TCC of the renal pelvis and ureter. Statistical analysis revealed close associations of both PI and MVD with tumor stage and of AI with tumor grade. Our study demonstrated a strong relationship between PI and MVD, but did not show associations of AI with PI or MVD in TCC of the renal pelvis and ureter. CONCLUSIONS: It is suggested that the high activity of tumor cell proliferation with rich neovascularization may be related to the high malignant potential of the cancer, and evaluation of cell proliferation combined with angiogenesis may be useful in predicting the progression of the renal pelvic and ureteral TCC.

Frequency and distribution of TUNEL-positive neurons in brains of dementia with Lewy bodies: comparison with those in brains of Alzheimer's disease.

The present study investigated the frequency and distribution of TUNEL-positive neurons in brains of dementia with Lewy bodies (DLB) in comparison with those in brains of Alzheimer's disease (AD), Down syndrome (DS) and non-demented elderly persons. In DLB brains, TUNEL-positive neurons were increased in frequency compared with those in non-demented elderly brains, and showed a distribution similar to those in AD and DS brains. DLB cases with TUNEL-positive neurons showing severe Lewy pathology were all neocortical type, while DLB cases of the limbic type showing mild Lewy pathology did not demonstrate TUNEL-positive neurons. In addition, we investigated the relationships between TUNEL-positive neurons and pathological hallmarks of DLB or AD brains. TUNEL-positive neurons had no Lewy bodies or neurofibrillary tangles, and were not located within amyloid deposits. These findings suggest that neuronal damage showing DNA fragmentations occurs in DLB brains as well as in AD and DS brains, and that it is accelerated by progression of Lewy pathology as well as Alzheimer pathology, although it is not directly related to their pathological hallmarks.

Antibody-regulated neurotoxic function of cell-surface beta-amyloid precursor protein.

APP is a transmembrane precursor of beta-amyloid, and its mutations cause early-onset familial Alzheimer's disease. We report a toxic function of normal wild-type APP (wtAPP). Treatment of neuronal F11 cells, immortalized embryonic day 13 neurons, overexpressing wtAPP with anti-APP antibodies caused death. Death was not induced by antibody in parental F11 cells. Death by antibody occurred through cell-surface APP, not through secreted APP, in a pertussis toxin-sensitive manner and was typical apoptosis, not observed in primary astrocytes or glioma cells overexpressing wtAPP, but observed in primary cortical neurons. Cell-surface APP thus performs a toxic function as an extracellularly controllable regulator of neuronal death. This study provides a novel insight into the normal and pathological functions of cell-surface wtAPP.

Loss of dystrophin and some dystrophin-associated proteins with concomitant signs of apoptosis in rat leg muscle overworked in extension.

This study investigated the basis for the high severity of damage to skeletal muscle due to eccentric exercise, i.e., to muscles generating force while lengthened. Fast and slow rat leg muscles maintained in an extended position were examined after 2-24 h of continuous stimulation. The treatment caused the injury to some regions of both muscles. Within the better preserved parts of the muscles, i.e., those without signs of necrotic processes, dystrophin, spectrin, and some of the dystrophin-associated proteins (beta-dystroglycan, alpha-sarcoglycan, and gamma-sarcoglycan) disappeared from sarcolemma of many fibers. The reduction or loss of dystrophin from the sarcolemma was more evident than that of other proteins examined, with sarcoglycans apparently being the most preserved. Several muscle fibers devoid of dystrophin contained apoptotic nuclei. Simultaneously, Bax, Bcl-2 and caspase-3 proteins appeared in many fibers. Our results indicate that a normal muscle overworking in an extended position undergoes the loss of several membrane skeletal proteins because of the excessive stress to the membrane cytoskeleton, which can lead to fiber death by either apoptosis or necrosis. This experimental model may represent a good model for mimicking the pathogenetic events in several muscular dystrophies.

DNA fragmentation, gliosis and histological hallmarks of Alzheimer's disease.

The extent of DNA fragmentation analysed using the TUNEL technique was evaluated in post-mortem human brain tissue. Twenty-four patients with clinical and histopathological diagnosis of Alzheimer's disease (AD) and a short post-mortem delay were analysed. We report an increase in the count of TUNEL-labelled cells as the pathology of AD intensifies. Our results point out a significant correlation between neurofibrillary tangle and senile/neuritic plaque score and TUNEL-labelled cells. Patients with two copies of apolipoprotein (Apo) Eepsilon4 allele had highest number of histopathological hallmarks lesions of AD, whereas the ApoE genotype did not significantly influence the density of TUNEL-positive cells. No significant correlation was found between beta-amyloid protein load and TUNEL-labelled cells. There was no relationship between the age at death, age at onset, extent of astrogliosis or microgliosis and TUNEL-labelled cells in our material.

Age-related changes of cornu ammonis 1 pyramidal neurons in gerbil transient ischemia.

This study reports that postischemic apoptotic cell death of the hippocampal cornu ammonis (CA) 1 neurons is delayed in aged gerbils. Age-related changes in the process of CA1 neuronal death following transient ischemia was studied. Two groups of Mongolian gerbils were used in this study, which compared adult (4-month-old) and aged (24-month-old) animals by hematoxylin-eosin stain, in situ nick-end labeling (TUNEL method) and electron microscopy. In the process of neuronal death, neuronal loss of the aged group was histologically less severe than that of the adult group. TUNEL-positive cells were found on days 3-5 after ischemia in the adult group, while they were still found on day 7 in the aged group. The apoptotic process of the aged group was delayed compared to the adult group. Furthermore, lipofuscin was ultrastructurally observed inside the apoptotic body 5 days after ischemia in CA1 pyramidal neurons of the aged group. It is likely that colocalization of lysosomal enzyme cathepsin D with lipofuscin might be associated with the age-related alteration of lysosomal system in the neurons. Altogether these data suggest that age-related lysosomal changes might affect the apoptotic cascade process in postischemic CA1 neurons.