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Terminal deoxynucleotidyl transferase (TdT) has been localized in both the nuclear and cytoplasmic compartments of lymphoblastic lymphomas when such tumors have been frozen for cryotomy and subsequently, formalin fixed and processed to paraffin. This cryotomy-induced antigen diffusion (CIAD) occurs with several different sources of anti-TdT antisera and was not observed in nonlymphoblastic tumors. CIAD was also not observed with other nuclear antigens including BRG1, Pax5, Ki67, BCL6, INI-1, myogenin, MyoD1, and Phox2B. TdT CIAD seems to be specific for lymphoblastic lymphomas. CIAD is another preanalytical artifact that needs to be considered in immunohistochemistry.

Background: Microwave therapy is a minimal invasive procedure and has been employed in clinical practice for the treatment of various types of cancers. However, its therapeutic application in non-small-cell lung cancer and the underlying mechanism remains to be investigated. This study aimed to investigate its effect on Lewis lung carcinoma (LLC) tumor in vivo. Methods: Fifty LLC tumor-bearing C57BL/6 mice were adopted to assess the effect of microwave radiation on the growth and apoptosis of LLC tumor in vivo. These mice were randomly assigned to 10 groups with 5 mice in each group. Five groups were treated by single pulse microwave at different doses for different time, and the other five groups were radiated by multiple-pulse treatment of a single dose. Apoptosis of cancer cells was determined by terminal deoxynucleotidyl transferase dUTP nick-end labeling assay. Western blotting was applied to detect the expression of proteins. Results: Single pulse of microwave radiation for 5 min had little effect on the mice. Only 15-min microwave radiation at 30 mW/cm2 significantly increased the mice body temperature (2.20 ± 0.82)°C as compared with the other groups (0.78 ± 0.29 °C, 1.24 ± 0.52 °C, 0.78 ± 0.42 °C, respectively), but it did not affect the apoptosis of LLC tumor cells significantly. Continous microwave radiation exposure, single dose microwave radiation once per day for up to seven days, inhibited cell division and induced apoptosis of LLC tumor cells in a dose- and duration-dependent manner. It upregulated the protein levels of p53, Caspase 3, Bax and downregulated Bcl-2 protein. Conclusions: Multiple exposures of LLC-bearing mice to microwave radiation effectively induced tumor cell apoptosis at least partly by upregulating proapoptotic proteins and downregulating antiapoptotic proteins. Continuous radiation at low microwave intensity for a short time per day is promising in treating non-small-cell lung cancer. Key words: Apoptosis; Lewis Lung Carcinoma Cells; Microwave Radiation; Non-small-cell Lung Cancer

Spontaneous remission of untreated pediatric leukemia is an extremely rare occurrence. The underlying mechanism may be because of an immune-mediated process or increased cortisol production during stress or infection. We describe a rare case of terminal deoxynucleotidyl transferase negative B-acute lymphoblastic leukemia with concurrent infection that went into remission without treatment with chemotherapy or corticosteroids. Though B-acute lymphoblastic leukemia can rarely go into spontaneous remission, these patients require close follow-up as most patients will eventually develop recurrence.

Estrogen plays an important role in cognitive function, including attention, learning, and memory, and affects the structure and function of brain areas. We investigated the effects of combined exercise on memory deficits induced by ovariectomy (OVX) in relation to cell proliferation and apoptosis in the hippocampus.Rats were randomly divided into four groups: sham, sham and exercise, OVX, and OVX and exercise. Rats in combined exercise groups were subjected to 3 days of resistance training and 3 days of running (for a total of 6 d/wk) for eight consecutive weeks. Rats were tested in step-down avoidance task and Morris water maze task to verify the effects of OVX on short-term and spatial working memory.In the present study, the number of BrdU-positive and doublecortin-positive cells and expression of brain-derived neurotrophic factor, TrkB, and Bcl-2 decreased; expression of Bax and the number of caspase-3-positive and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive cells increased; and short-term and spatial working memory decreased in the OVX group compared with the sham group. Conversely, when the combined exercise group was compared with the OVX group, the number of BrdU-positive and doublecortin-positive cells and expression of brain-derived neurotrophic factor, TrkB, and Bcl-2 increased; expression of Bax and the number of caspase-3-positive and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive cells decreased; and short-term and spatial working memory increased.Combined exercise increases cell proliferation and inhibits apoptosis in the hippocampus and improves cognitive function despite estrogen deficiency.

T-lymphoblastic lymphoma is an aggressive neoplasm requiring prompt clinical treatment. Conversely, indolent T-lymphoblastic proliferation mimics T-lymphoblastic lymphoma but consists of a proliferation of non-neoplastic TdT+ T cells, requiring no treatment. Recently, we identified several cases of indolent T-lymphoblastic proliferations in extrathymic lymphoid tissues: 1 in a patient suffering from Castleman disease (CD) associated with a follicular dendritic cell sarcoma/tumor, 1 in a patient with a history of angioimmunoblastic T-cell lymphoma (AITL), and 1 in association with acinic cell carcinoma. Interestingly, in the case of the patient with a history of AITL, these TdT+ T cells were seen in multiple anatomic sites over the span of 5 years. Here we review these 3 cases and extend our findings by demonstrating that TdT+ T-lymphoblastic populations are increased in lymph nodes of patients with CD (P=0.011), CD in association with follicular dendritic cell tumors, and AITL (P

The purpose of this study is to investigate the mixed lineage leukemia gene (MLL, located on chromosome 11q23) expression in T acute lymphoblastic leukemia (T-ALL)/lymphoblastic lymphoma (LBL) and its relationship to prognosis. Fifty cases of T-ALL/LBL with clinical data were selected from the Shanxi Cancer Hospital in China. The immunohistochemical EnVision method was used for the expression of CD3, CD7, CD10, CD20, CD23, CD43, CD45RO, CD99, terminal deoxynucleotidyl transferase, myeloperoxidase and ki67. Fluorescent in situ hybridization for MLL gene expression was performed on paraffin-embedded tissue. Among the 50 cases of T-ALL/LBL, the percentages of tumor cells expressing terminal deoxynucleotidyl transferase, CD99, CD3, CD7, CD10, CD43, and CD45RO were 92.0%, 96.0%, 72.0%, 92.0%, 34%, 60.0%, and 40.0%, respectively, whereas myeloperoxidase, CD20, and CD23 were all negative. A level of Ki67 expression >80% was found in 18 cases and ≤80% in 32 cases. The period of follow-up ranged from 1 to 108 months. The overall survival rate was 35.8%, with a median survival time of 330 days. Breakage of 11q23 was detected in 8 (16.00%) and amplification in 14 (28.00%) of the 50 cases. The rate of amplification in stage III-IV was higher than that in stage I-II (P

Objective To ascertain whether Premarin improves spinal cord injury outcomes in male rats by stimulating both angiogenesis and neurogenesis. Design Chi Mei Medical Center research laboratory. Subjects Male Sprague-Dawley rats 240-258 g. Interventions Anesthetized rats, after the onset of spinal cord injury, were divided into two groups and given the vehicle solution (1 mL/kg of body weight) or Premarin (1 mg/kg of body weight). Saline or Premarin solutions were administered intravenously and immediately after spinal cord injury. Measurements and main results Premarin (an estrogen sulfate) causes attenuation of spinal cord injury-induced spinal cord infarction and hind limb locomotor dysfunction. Spinal cord injury-induced apoptosis as well as activated inflammation was also significantly Premarin-reduced. In injured spinal cord, angiogenesis, neurogenesis, and production of an antiinflammatory cytokine were all Premarin therapy-promoted. Conclusions Our results indicate that Premarin therapy may protect against spinal cord apoptosis after spinal cord injury through mechanisms stimulating both angiogenesis and neurogenesis in male rats.

Background: The purpose of this study was to examine the role of erythropoietin in retinal ischemic preconditioning (IPC). Methods: Rats were subjected to retinal ischemia after IPC. Electroretinography assessed functional recovery after ischemia; retinal sections were examined to determine loss of retinal ganglion cells, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling was used to assess apoptosis. Levels of downstream mediators were measured in retinal homogenates by Western blotting. To assess the involvement of erythropoietin in IPC, Western blotting was used to measure levels of erythropoietin and its receptor (EPO-R) in retinal homogenates after IPC. To examine erythropoietin's role in IPC, the impact of blocking erythropoietin via intravitreal injection of soluble EPO-R (sEPO-R) before IPC was studied. Results: Erythropoietin levels did not change after IPC, but EPO-R increased. Intravitreal injection of sEPO-R significantly attenuated both the functional and histologic neuroprotection produced by IPC in comparison to control injection of denatured sEPO-R. Apoptotic damage after ischemia was enhanced in the sEPO-R-treated retinas as indicated by fluorescent terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling. Phosphorylated extracellular-signal-regulated kinase and heat shock protein 27, but not protein kinase B, upregulated in denatured sEPO-R-treated retinae, were attenuated in eyes injected with sEPO-R. Conclusions: These results indicate that EPO-R upregulation is a critical component of the functional, histologic, and anti-apoptotic protective effect of IPC on ischemia in the retina and that several downstream effectors may be involved in the neuroprotective actions of erythropoietin.

Accurate bone marrow (BM) blast counts (BCs) are essential for diagnosis (dx) of myelodysplasia (MDS), MDS/myeloproliferative (MDS/MPD) disease, or acute myeloid leukemia (AML), and may be difficult in hemodiluted bone marrow aspirates (BMAs). Erythroid precursors (EPs) may be indistinguishable from myeloblasts in BM sections (aspirate clots/cores). We compare the usefulness of immunohistochemistry (IHC) [ie, CD34, CD117, myeloperoxidase (MPO), Hemoglobin A1 (HbA1), and terminal deoxynucleotidyl transferase (TdT)] of BM sections (IHC-BM) with BMA, bone marrow touch preparation (BMTP), and flow cytometry (FC) BCs.The initial BC (48), percentage (%) of Eps (38) (both based on initial 100 to 600-cell counts), and FC expressions of CD34, CD117, Glycophorin A(GLY A), and TdT (44) were tabulated from 50 BMs (MDS, MDS/MPD, or AML). BMAs (48) and BMTPs (25) subsequently received 500-cell counts. IHC-BM was performed (45:formalin, 5:B5-fixed) [CD34 (46), CD117 (45), HbA1 (45), TdT (42), and MPO (45)].Retrospective BMA BCs revealed a 31% (15/48) discrepant rate between the original/retrospective BMA BCs; 80% revealed an underestimated initial BC. There was a 28% discordance rate between the retrospective BMA and BMTP reviews; 77% showed a higher BMTP BC. IHC showed significantly higher BCs in 19% (9/47), resulting in a different dx (5). However, CD34 and CD117 IHCS revealed lower BCs in 38% and 48%, respectively. The CD34 IHC results were primarily due to CD34-negative blasts by FC. The CD117 IHC results were largely unexplained. EPs were CD34 and CD117-negative.(1) Evaluation for MDS/AML requires 500-cell counts of BMAs and/or BMTPs. (2) CD34 and/or CD117 blasts by FC indicate IHC-BM may increase BC accuracy. (3) CD34 is more reliable than CD117 by IHC; however, in combination, they are most reliable and should be performed on BM clots/cores due to variable reactivity.

Primary lymphoma of the central nervous system (PCNSL) is extremely rare, especially in childhood. A 9-year-old Japanese boy was diagnosed as having precursor-B cell-type lymphoblastic lymphoma, based on morphologic and immunocytochemical analysis of mononuclear cells in the cerebrospinal fluid and a positive reaction for terminal deoxynucleotidyl transferase (TdT), CD19, CD79a, and CD179b. After seven courses of chemotherapy and craniospinal radiotherapy, the patient is alive, well, and in continuous complete remission. Despite its rarity, PCNSL should be included in the differential diagnosis in the presence of symptoms of increased intracranial pressure and/or unusual imaging findings of the brain.

We report 2 cases of microscopic-sized thymoma, which probably represents the earliest phase of thymoma development. The 2 patients presented with pure red cell aplasia and myasthenia gravis, respectively. The thymectomy specimens did not reveal tumor on gross examination, but histologically each contained small thymomas measuring 5 mm and 7 mm in largest dimension, respectively. One of the tumors was unencapsulated and involved a single lobule only, and the other was encapsulated and comprised two lobules. The tumors consisted of ovoid epithelial cells with pale nuclei and distinct nucleoli, scattered in a background of small lymphocytes. Foci of medullary differentiation and perivascular space were identified in the 2 cases, respectively. The lymphocytes were confirmed to be immature T cells on immunohistochemical studies (CD3+, TdT+). Except for the microscopic size, the morphology of the two tumors conforms to conventional type B1/B2 and type B2 thymoma, respectively. We propose calling such incidental small tumor "microthymoma" to distinguish it from the so-called microscopic thymoma, which is composed of small thymic epithelial nests and probably more appropriately termed "nodular hyperplasia" of the thymic epithelium.

Terminal deoxynucleotidyl transferase (TdT), a template-independent DNA polymerase, contributes to antigen receptor diversity in lymphocytes. Using in situ hybridization, we found that tdt is expressed within neurons of the adult mouse brain. tdt mRNA was localized within pyramidal neurons in the hippocampus, granule and polymorphic cells in the dentate gyrus, Purkinje neurons in the cerebellum, and cortical cells. Increased levels of tdt mRNA in the hippocampus, neocortex, and cerebellum were associated with rearing C57BL/6 mice, but not DBA/2 mice, in enriched environments. Unlike wild types (WT), tdt (-/-) mice did not show improvement in spatial learning and memory as a result of rearing in enriched environments. These results suggest that tdt may be involved in learning and memory saving.

The classification of blastic or blastoid natural killer (NK)-cell lymphoma is controversial. Reports of primary cutaneous blastic CD56+ NK-cell lymphoma are rare, which necessitates further clinicopathologic definition of this type of lymphoma. Most CD56+ lymphomas display angiocentric histologic features, especially in Asian patients, and these are mostly associated with the presence of Epstein-Barr virus (EBV) genome and with an aggressive clinical course. We report on a young woman with a primary cutaneous blastic NK lymphoma which showed no angiocentric features but showed an unusual immunophenotype; CD56+, TdT+, CD4+, EBV-, and germline configuration of T-cell receptor gene. This unusual lymphoblastic lymphoma seems to have an immature or progenitor NK cell lineage.

The isoprenoids geranylgeraniol and farnesol are required for lipid modification of several important cellular proteins. In this study the effect of perillyl alcohol (PA), an inhibitor of geranylgeranyl transferase, was examined on proliferation and apoptosis of immortalized human vascular smooth muscle cells (HVSMCs) derived from saphenous vein. PA induced a time- and concentration-dependent inhibition of cell proliferation over 3 days and 48 h exposure to PA inhibited [methyl-3H-thymidine incorporation induced by 10% fetal calf serum in a concentration-dependent manner. Flow cytometry analysis indicated that PA reduced the proportion of cells in S phase and increased apoptosis. PA-induced apoptosis was confirmed by terminal deoxynucleotidyl transferase (TdT) reagent-based immunocytochemistry. In contrast, the selective inhibitors of farnesyl transferase B581 and BZA-5B were without effect. In view of the proposed role of proliferation and apoptosis in myointimal hyperplasia, inhibitors of geranylgeranyl transferases may have therapeutic potential in cardiovascular disease.

To define the functions of retinoids and their receptors in insulin secretion, we tested the effects of all-trans-retinoic acid (ATRA) and retinoic acid receptor (RAR) expression on cell growth, differentiation, and secretion using insulin-secreting RINm5F cells. Wild-type cells with a low abundance of mRNA for RAR beta were transfected with RAR beta or chloramphenicol acetyltransferase (CAT control). Cells were cultured for 2-7 days in media without (A-def) or with ATRA, 1, 10, 100, and 1,000 nM. At day 2 of culture, ATRA stimulated insulin release in wild-type and transfected cells, and this effect was dose dependent. At 7 days, ATRA stimulated insulin secretion from wild-type cells twofold at glucose concentrations of 0.5 mM (A-def, 5.1 +/- 0.27; ATRA, 1,000 nM, 10.5 +/- 1.43 ng/10(6) cells) and at 11.0 mM (A-def, 6.9 +/- 0.24; ATRA, 1,000 nM, 13.6 +/- 1.86 ng/10(6) cells). The cellular insulin content was increased about threefold (A-def, 39.2 +/- 2.95; ATRA, 1,000 nM, 118 +/- 8.54 ng/10(6) cells). ATRA inhibited growth of wild-type cells as early as 3 days, and this effect was dose dependent. Whereas in the absence of ATRA, the cell number increased over fivefold between day 3 and day 5, ATRA, 1,000 nM, inhibited cell growth completely. ATRA, 1,000 nM, increased apoptotic RINm5F cells (day 3 A-def, 0.53 +/- 0.27% of total cells, and ATRA, 2.30 +/- 1.44; day 5 A-def, 0.38 +/- 0.23, and ATRA, 2.14 +/- 0.59; day 7 A-def, 0.90 +/- 0.29, and ATRA, 6.02 +/- 1.64). RAR beta-transfected cells showed overexpression of mRNA to RAR beta and dose-dependent inhibition of growth, with almost-complete inhibition at ATRA concentrations as low as 100 nM. Overexpression of RAR beta increased insulin secretion at ATRA, 100-1,000 nM. In summary, ATRA increased the insulin secretion and content of RINm5F cells, while inhibiting growth and increasing apoptosis. Increased expression of RAR beta facilitated these effects on growth and secretion. These findings may reflect the known effect of ATRA on differentiation of cells and mediation through RAR beta.

To seek evidence that tumor regression in neuroblastoma might result from massive apoptosis, we investigated tumor cell death in 39 neuroblastomas. Characteristic histologic features of apoptosis, condensed nuclear fragments and eosinophilic cytoplasm, were observed in all specimens. A ladder of DNA fragments induced by apoptosis was demonstrated by means of DNA agarose gel electrophoresis in 18 of the 19 tumors examined. In situ DNA nick end labeling (TUNEL) stained the nuclei with DNA fragmentation in 16 of 39 neuroblastomas. The TUNEL -positive cells were distributed in a scattered fashion in 10 tumors. In the remaining six tumors, they were densely located around nonviable areas of calcifications, where karyorrhectic or pyknotic cells were frequently observed. Five of six patients with such tumors were under 12 months of age, but there was no significant difference between the two groups in the patient age, origin of the primary lesion, or tumor stage. Biological features, including histology. DNA ploidy, and N-myc amplification, were not significantly different . Double fluorescent staining for bcl-2 oncoprotein and TUNEL showed that bcl-2 oncoprotein was expressed in the cytoplasm of tumor cells that were negative for TUNEL staining. This accumulated evidence suggests that massive apoptosis of tumor cells occurs in some neuroblastomas and may be related to tumor regression, whereas inhibition of apoptosis by bcl-2 oncoprotein expression might be associated with the tumorigenesis of neuroblastomas, as reported in our previous study.

Xanthine oxidase (XO) activity and hydroxyl radical (.OH) formation are widely proposed mediators of renal reperfusion injury, potentially altering the severity of, and recovery from, postischemic acute renal failure. The goal of this study was to ascertain whether combination XO inhibitor (oxypurinol) and .OH scavenger (Na benzoate) therapy, given at the time of renal ischemia, alters the extent of: (1) tubular necrosis and filtration failure; (2) DNA fragmentation/apoptosis (assessed in situ by terminal deoxynucleotidyl transferase reactivity); (3) early tubular regenerative responses (proliferating cell nuclear antigen expression; (3H)thymidine incorporation); and (4) the rate and/or degree of functional and morphologic repair. The effects of XO inhibition, .OH scavengers, and "catalytic" iron (FeSO4) on human proximal tubular cell proliferation in vitro were also assessed with a newly established cell line (HK-2). Male Sprague-Dawley rats were subjected to 35 min of bilateral renal arterial occlusion with or without oxypurinol/benzoate therapy. These agents did not alter the extent of tubular necrosis or filtration failure, proliferating cell nuclear antigen expression or thymidine incorporation, or the rate/extent of renal functional/morphologic repair. DNA fragmentation did not precede tubular necrosis, and it was unaffected by antioxidant therapy. By 5 days postischemia, both treatment groups demonstrated regenerating epithelial fronds that protruded into the lumina. These structures contained terminal deoxynucleotidyl transferase-reactive, but morphologically intact, cells, suggesting the presence of apoptosis. Oxypurinol and .OH scavengers (benzoate; dimethylthiourea) suppressed in vitro tubular cell proliferation; conversely, catalytic Fe had a growth-stimulatory effect. These results suggest that: (1) XO inhibition/.OH scavenger therapy has no discernible net effect on postischemic acute renal failure; (2) DNA fragmentation does not precede tubular necrosis, suggesting that it is not a primary mediator of ischemic cell death; and (3) antioxidants can be antiproliferative for human tubular cells, possibly mitigating their potential beneficial effects.

✓ This study explores the effects of infusion of nerve growth factor (NGF) on behavioral outcome and cell death in the septal region using the clinically relevant model of fluid-percussion brain injury in the rat. Animals were subjected to fluid-percussion brain injury and 24 hours later a miniosmotic pump was implanted to infuse NGF (12 animals) or vehicle (12 animals) directly into the region of maximum injury for 2 weeks. Four weeks postinjury the animals were tested for cognitive function using a Morris Water Maze paradigm. Neurological motor function was evaluated over a 4-week postinjury period. The rats receiving NGF infusions had significantly higher memory scores than vehicle-treated animals. Examination of the cholinergic neurons in the medial septal region using choline acetyltransferase immunohistochemistry demonstrated significant cell loss after injury. Infusion of NGF significantly attenuated loss of these cholinergic neurons. A second group of animals was subjected to fluid-percussion brain injury alone (23 rats) or injury followed by NGF infusion (18 rats). These animals were killed between 24 hours and 2 weeks postinjury and the septal region was examined for the presence of apoptotic cells using the terminal deoxynucleotidyl transferase—mediated biotinylated-deoxyuridinetriphosphate nick-end labeling technique. Apoptotic cells were identified as early as 24 hours postinjury; their numbers peaked at 4 and 7 days, and then declined by 14 days. The NGF-treated animals had some apoptotic cells; however, even at 7 days there were significantly fewer of these cells. No significant motor differences were observed between the NGF- and vehicle-treated groups. These data indicate that NGF administration beginning 24 hours after fluid-percussion brain injury has a beneficial effect on cognition and results in sparing of cholinergic septal neurons. These improvements persist after cessation of NGF administration. The beneficial effects of NGF may be related to its ability to attenuate traumatically induced apoptotic cell death.