Bromodeoxyuridine Labeling Index as an Indicator of Early Tumor Response to Preoperative Radiotherapy in Patients with Rectal Cancer

Purpose Assessment of tumor proliferation rate using Bromodeoxyuridine labeling index (BrdUrdLI) as a possible predictor of rectal cancer response to preoperative radiotherapy (RT). Methods and material Ninety-two patients were qualified either to short RT (5 Gy/fraction/5 days) and surgery about 1 week after RT (schedule I), or to short RT and 4–5 weeks interval before surgery (schedule II). Tumor samples were taken twice from each patient: before RT and at the time of surgery. The samples were incubated with BrdUrd for 1 h at 37°C, and the BrdUrdLI was calculated as a percentage of BrdUrd-labeled cells. Results Thirty-eight patients were treated according to schedule I and 54 patients according to schedule II. Mean BrdUrdLI before RT was 8.5% and its value did not differ between the patients in the two compared groups. After RT tumors showed statistically significant growth inhibition (reduction of BrdUrdLI). As the pretreatment BrdUrd LI was not predictive for early clinical and pathologic tumor response, prognostic role of the ratio of BrdUrdLI after to BrdUrdLI before RT was considered. The ratios were calculated separately for fast (BrdUrd LI > 8.5%) and slowly (BrdUrd LI ≤ 8.5%) proliferating tumors and correlated with overall treatment time (OTT, i.e., time from the first day of RT to surgery). One month after RT, accelerated proliferation was observed only in slowly proliferating tumors. Conclusions Pretreatment BrdUrdLI was not predictive for early clinical and pathologic tumor response. The ratio after/before RT BrdUrdLI was correlated to inhibition of proliferation in responsive tumors. The paper was presented at ECCO 13, October 30 to November 03, 2005 in Paris, France     

左旋肉毒碱对人成骨细胞增殖和凋亡的影响

目的观察左旋肉毒碱(L-carnitine,L-C)对人成骨细胞增殖和凋亡的作用。方法以3H掺入法(3H-TdR)测定左旋肉毒碱对人成骨细胞增殖的影响;以吖啶橙/溴化乙啶染色和细胞凋亡ELISA(酶联免疫吸附法)试剂盒测定左旋肉毒碱对人成骨细胞凋亡的影响;以免疫印记法检测左旋肉毒碱干预前后活化型Caspase-3、-9蛋白质表达水平的改变。结果10~100mmol/L左旋肉毒碱能促进人成骨细胞增殖。吖啶橙/溴乙啶染色法示100mmol/L左旋肉毒碱干预使人成骨细胞凋亡细胞明显减少。ELISA结果示10~100mmol/L左旋肉毒碱干预可显著抑制人成骨细胞凋亡。10~100mmol/L左旋肉毒碱干预可抑制无血清饥饿诱导的人成骨细胞caspase-3、9活化增加。结论左旋肉毒碱促进人成骨细胞增殖,并通过下调Caspase-3、-9的活化抑制人成骨细胞凋亡。     

苦参碱对人HT1080细胞系增殖抑制的体外研究

目的：研究苦参碱（Matrine)对人成纤维肉瘤（HT1080）细胞系生长的抑制效应，并探讨其作用机制。方法：用不同浓度的Mat加入体外培养HT1080细胞中，观察加药后细胞生长数量及其形态的变化；用MTT法检测Mat的细胞毒作用。结果：Mat明显抑制HT1080细胞生长；IC50值为350μg/ml；其抑制率与药物浓度呈剂量依赖关系，结论：Mat能有效抑制HT1080细胞的增殖。     

A study on OX39, a murine anti-rat interleukin 2 receptor antibody

OX39, a murine IgG1 monoclonal antibody (MoAb) that recognizes the 55 kDa alpha chain of the rat interleukin 2 receptor (R-IL2), was studied in vitro for its ability to interfere with IL2 binding and IL2-induced proliferation on rat concanavalin A (ConA) blasts and in vivo in a model of rat heart allografts. In vitro studies indicated that OX39 MoAb interacts with a single class of sites on the alpha chain of the rat R-IL2 with a high affinity (K_D=0.8 nm) and competes with IL2 binding on this chain (K_I=0.53 nm). In contrast, OX39 MoAb was found to be 10–20 times less efficient in competing with IL2 binding to the high-affinity R-IL2 (K_I10 nm). It is proposed that the epitope recognized by OX39 on the alpha chain (low-affinity R-IL2) is modified on (or buried in) the high-affinity R-IL2 configuration. Accordingly, OX39 was found to be a weak inhibitor in vitro on IL2-induced proliferation and in vivo on allograft rejection. Allograft survival was unaffected by doses of OX39 of 20 and 50 g/rat for 9 days; only a borderline effect was noted when doses as high as 250 g/rat were used. A significant, but restricted, effect of OX39 could be further detected when combined with low doses of cyclosporine A (1.5 mg/kg), which were ineffective by themselves. Together, our data suggest that in order to be efficient in vivo, anti-R-IL2 MoAbs must bind with high affinity to epitopes involved in the high-affinity IL2 binding site.     

Dietary restriction: effects of short-term fasting on protein uptake and cell death/proliferation in the rat liver

Dietary restriction (DR) is known to prolong life in laboratory animals. Intermittent (alternate-day) fasting or short-term repeated fasting has also been reported to increase the life span of animals. In the present study, we investigated the changes or induction of abnormalities of protein metabolism in rats during fasting, and measured asialoglycoprotein uptake and cell death/proliferation in the liver of rats receiving fasting and refeeding. In the results, liver weight decreased significantly after 48 h of fasting and increased during the refeeding period, returning to the pre-fasting level by 12 h of refeeding. Cell death, determined by single stranded DNA (ssDNA) staining method, increased during the fasting period, and returned to the pre-fasting level during the refeeding period. Cell proliferation, determined using antibodies (Ab) against proliferating cell nuclear antigen, decreased during the fasting period, and increased during the refeeding period. Changes in cell death and cell proliferation were inversely related. However, there was no significant difference in asialoglycoprotein uptake by the whole liver between the ad libitum (AL)-fed rats and 48 h fasted rats. Thus, neither the changes in liver weight nor cell death/proliferation affected asialoglycoprotein uptake on a living body. These results suggest that episodes of 48 h fasting do not induce protein metabolism abnormalities in the liver.     

The proliferative capacities of popliteal lymph node lymphocytes and of their functionally distinct T-cell subpopulations from young-adult and old mice

Young adult and old mice were immunized by footpad injection of dinitrophenyl-conjugated bovine gamma-globulin (DNP-BGG) in complete Freund's adjuvant. A comparison of lymph node weight and total number of nucleated cells per lymph node as a function of time after antigen injection demonstrated a significantly greater absolute increase in lymph node weight and peak number of nucleated cells per lymph node in young-adult than in old animals. However, as judged by this increase in total nucleated cells, other than being delayed in old mice, the magnitude of these in situ proliferative responses appeared comparable for young-adult and old mice. That is, the antigen-stimulated to non-stimulated cell ratio did not differ significantly between young-adult and old animals. This was because lymph nodes from old animals prior to antigen injection always weighed less and had fewer numbers of nucleated cells compared with young-adult animals. Therefore, the in vitro cellular proliferative response of three T-cell-enriched lymphocyte subpopulations from young-adult and old mice was further characterized. This was done by measuring [3H]thymidine incorporation following antigen- (DNP-BGG)- or mitogen-[phytohemagglutinin (PHA) or Concanavalin A (Con A)]-induced proliferation and assessing their quantitative and/or qualitative requirements for macrophages. In contrast to the markedly reduced proliferation of the two T-cell subpopulations from popliteal lymph nodes which respond to PHA and Con A in old animals primed 21-days earlier with DNP-BGG, antigen-induced in vitro cellular proliferation of the small T-cell subset in old mice specifically responsive to the immunizing antigen DNP-BGG always responded as well as, if not better than, cells from young-adult mice.     

Glomerular expression of cell-cycle-regulatory proteins in human crescentic glomerulonephritis

To elucidate the mechanism underlying crescentic formation, we assessed the phenotypic characterization and cell-cycle protein expression in human crescentic glomerulonephritis (CRGN). Kidney tissue specimens taken from CRGN patients (10 patients with pauci-immune type rapidly progressive glomerulonephritis (RPGN), 2 patients with Henoch-Schönlein purpura nephritis, and 1 patient with IgA nephropathy) were examined immunohistochemically. Most of the cellular components of the crescents expressed cytokeratin, whereas few cells expressed PHM-5. CD68-positive cells were minor components of cellular crescents, indicating that the major principal cellular component of the crescents is made up of cells with the parietal glomerular epithelial cell (PEC) phenotype. Additionally, serial section analysis revealed that Ki-67-positive cells in the crescents were frequently cyclin-A positive and Bcl-2 positive, but seldom cyclin-B₁ positive. Moreover, the expression of cyclin-dependent kinase inhibitor p27^Kip1 was low in the cellular crescents, despite being exclusively positive in podocytes within the same section. We concluded that the major component of the cellular crescents is made up of PECs and that apparent expression of cyclins and Bcl-2 and restrained expression of p27^Kip1 may be synergistically associated with the development of cellular crescents in human CRGN.     

Apoptosis, proliferation and gene expression patterns in mouse developing tongue

The Fgf/Fgfr (Fgf receptor) and Bmp signal pathways are critical for embryonic development and postnatal growth. In order to address their roles in tongue development, preliminary study of expression patterns of some important members in the two families, as well as of apoptosis and proliferation, were carried out in mouse developing tongue. Apoptosis in tongue is a very late event in embryogenesis, restricted to the upper layer of the epithelium whereas proliferation is very vigorous at the early stage of tongue development and remains active throughout embryogenesis. Bmp2, −4 and -5 were localized within the mesenchyme at the early embryonic stage of tongue development (E12 to E13), whereas Bmp3 and Bmp7 were mainly expressed in the epithelium. Most of these molecules were also seen in the tongue muscles at postnatal stages. Among Fgfr isoforms, Fgfr1c, −2b, and -2c were detected in embryogenesis with peak expression at E11 to E13. Fgfr1c and Fgfr2c were localized within the mesenchyme, while Fgfr2b was mainly expressed in the epithelium. High expression of Fgf7 and Fgf10 was also detected in the mesenchyme at the early embryonic stage of tongue development, corresponding to the Fgfr expression, suggesting that they are among the principal ligands functioning at the early embryonic expanding stage. Fgf2 was seen in the tongue muscles at the late embryonic and postnatal stages. These results suggest that Bmp and Fgf signalling regulates tongue development at multiple stages, possibly related to proliferation and differentiation.     

SRBC膜提取物及其对猪PBMNC激活作用的研究

胰酶水解绵羊红细胞（ＳＲＢＣ）膜得到粗提物，经离子交换层析部分纯化后，获得组分ＴＲＦ－Ⅲ（Ｔｒｙｐｓｉｎ－ｒｅｌｅａｓｅｄｆｒａｃｔｉｏｎⅢ）具有明显抑制ＳＲＢＣ与猪外周血淋巴细胞形成Ｅ玫瑰花的活性，花环形成率与ＴＲＦ－Ⅲ含量有明显的函数关系，电泳测定其分子量为６６ｋＤ。ＴＲＦ－Ⅲ单独作用尽管对ＰＢＭＮＣ的增殖无作用，但可增强ＰＨＡ刺激ＰＢＭＮＣ的增殖作用，ＴＲＦ－Ⅲ单独作用于ＰＢＭＮＣ可显著增强     

The small subset of CD56brightCD16- natural killer cells is selectively responsible for both cell proliferation and interferon-gamma production upon interaction with dendritic cells

The encounter of NK cells with dendritic cells (DC) undergoing maturation may result in the induction of NK cell proliferation. Whether such proliferation involves most NK cells or just a subset has yet to be determined. In the present study we analyzed the nature of such proliferating NK cells by combining carboxyfluorescein succinimidyl ester staining and double-fluorescence cytofluorimetric analysis. Freshly isolated peripheral blood NK cells cultured with LPS and immature DC underwent proliferation; however, proliferating cells were confined to a minor NK cell subset. This subset is characterized by the CD56(bright)CD16(-)NKG2A(+)KIR(-) surface phenotype (KIR, killer Ig-like receptor). This was further confirmed by the fact that, after cell sorting, only the CD56(bright) NK cells were able to proliferate in response to the DC stimulus, whereas the CD56(dull) were not. We also provide evidence that the CD56(bright) subset is the main source of IFN-gamma-producing NK cells, upon interaction with DC. The CD56(bright)CD16(-) NK cells express a panel of surface molecules including CD62L, CCR7 and CXCR3 that may allow their homing either to secondary lymphoid compartments or to inflamed tissues. This implies that, in vivo, the interactions between DC undergoing maturation and CD56(bright) NK cells may occur in different tissues and have different functional implications.