E2F decoy oligodeoxynucleotide ameliorates cartilage invasion by infiltrating synovium derived from rheumatoid arthritis

This study examined the ability of E2F decoy oligodeoxynucleotides (ODN) to inhibit proliferation of synovial fibroblasts derived from patients with rheumatoid arthritis (RA). The effect of E2F decoy ODN on cartilage invasion by RA synovium in a murine model of human RA was also investigated. E2F decoy ODN were introduced into synovial tissue and synovial fibroblasts derived from patients with RA using hemagglutinating virus of Japan (HVJ)-liposomes. The effect of E2F decoy ODN on synovial fibroblast proliferation was evaluated by MTT assay and by RT-PCR for the cell cycle regulatory genes proliferating-cell nuclear antigen (PCNA) and cyclin-dependent kinase 2 (cdk2). Changes in production of inflammatory mediators by RA synovial tissue following transfection with E2F decoy ODN were assessed by ELISA. Human cartilage and RA synovial tissue transfected with E2F decoy ODN were co-transplanted in severe combined immunodeficient (SCID) mice. After 4 weeks, the mice were sacrificed and the implants histologically examined for inhibition of cartilage damage by E2F decoy ODN. E2F decoy ODN resulted in significant inhibition of synovial fibroblast proliferation, corresponding with reduced expression of PCNA and cdk2 mRNA in synovial fibroblasts. The production of interleukin-1beta (IL-1beta), IL-6 and matrix metalloproteinase (MMP)-1 by synovial tissue was also significantly inhibited by the introduction of E2F decoy ODN. Further, in an in vivo model, cartilage that was co-implanted with RA synovial tissue transfected with E2F decoy ODN exhibited no invasive and progressive cartilage degradation. These data demonstrate that transfection of E2F decoy ODN prevents cartilage destruction by inhibition of synovial cell proliferation, and suggest that transfection of E2F decoy ODN may provide a useful therapeutic approach for the treatment of joint destruction in arthritis.     

Nestin expression in repopulating mesangial cells promotes their proliferation

We investigated whether the intermediate filament protein and neural stem cell marker nestin characterizes the glomerular progenitor/reserve cell population immigrating the glomerulus after mesangial cell (MC) injury in the rat (anti-Thy1 nephritis). Nestin expression was investigated by immunohistochemistry and real-time PCR during anti-Thy1 nephritis. Migration and proliferation assays were used to characterize the function of nestin in isolated MCs after nestin knockdown by siRNA. After MC injury during anti-Thy1 nephritis, glomerular nestin was transiently increased during the repopulation phase. At the peak of mesangial proliferation and expansion (day 5) most OX-7-positive MCs expressed nestin largely colocalizing with the activation marker alpha-smooth muscle actin and the proliferation marker PCNA. In contrast to a healthy, non-injured mesangium in vivo, MCs in culture are considered to be in an 'activated, injured state' and express nestin in a generalized distribution with condensed localization around the nucleus as well as intensive staining of cell protrusions such as filopodia. During cell cycle, the percentage of MCs with high nestin levels was increased during S- aupnd G2-phase. Blocking of nestin using specific siRNA resulted in inhibition of cell proliferation but not cell migration. In conclusion, nestin is constitutively expressed in podocytes, but is a marker for repopulating MCs after experimental MC injury in vivo. Nestin promotes MC proliferation in vitro, suggesting a supporting role for nestin during repair reaction.     

Role of differential and cell type-specific expression of cell cycle regulatory proteins in mediating progressive glomerular injury in human IgA nephropathy

The activities of cell cycle regulatory proteins have been reported to be associated with the development of pathological lesions in glomerulonephritis. To assess the cellular mechanisms underlying the mesangial cell proliferation and glomerulosclerosis in progressive human IgA nephropathy (IgAN), we examined the expression of E2F1, Rb, c-Myc, proliferating cell nuclear antigen (PCNA), cyclins (D1, E and A), cyclin-dependent kinase 2 (CDK2) and CDK inhibitors (p21(waf1), p27(kip1), 57(kip2) and p16(ink4a)) by immunohistochemistry in renal biopsy specimens. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) was also performed to detect the presence of apoptosis. In total, 51 cases of IgAN were categorized into four subgroups according to histological severity. A dramatic upregulation of E2F1 expression in mesangial cells was identified in proliferating glomeruli, which correlated well with the proliferation index. High endogenous expression of p27(kip1) and p57(kip2) by podocytes in normal glomeruli and glomeruli with minor lesions was observed to decrease in proliferating and sclerosing glomeruli; this pattern displayed a strong inverse correlation with the mean glomerulosclerosis score and the index of glomerular lesion. Increased apoptotic activity was identified in progressive glomerular lesions of advanced IgAN, which correlated with the proliferative activity in these lesions as assessed by total expression levels of PCNA and CDK2 in glomeruli, E2F1 expression levels in the mesangium, cyclin D1 expression levels in endothelium and the c-Myc glomerular staining score. Our results suggest that the onset and magnitude of mesangial cell proliferation and glomerulosclerosis is associated with the upregulation of E2F1 by mesangial cells and the downregulation of p27(kip1) and p57(kip2) by glomerular epithelial cells. The cell type-specific and coordinated regulation of proliferative and proapoptotic activities of cell cycle regulatory proteins may play an important role in mediating progressive glomerular injury in human IgAN.     

Expression of cell cycle regulators during smooth muscle cell proliferation after balloon catheter injury of rat artery

Intimal hyperplasia is defined as the abnormal migration and proliferation of vascular smooth muscle cells (VSMCs) with deposition of extracellular matrix. However, the cell cycle regulatory mechanisms of injury-induced VSMC proliferation are largely unknown. To examine the expression kinetics of cell cycle regulatory factors which is known to be worked positively or negatively, we used rat balloon injury model. Marked induction of proliferating cell nuclear antigen (PCNA), G1/S cyclin-dependent kinase (cdk2), and its regulatory subunit (cyclin E) occurred between 1 and 3 days after balloon arterial injury, and this was sustained for up to 7 days and then declined. However, the induction of the negative regulators, p21 and p27, occurred between 3 and 5 days of injury, peaked after 7 and 14 days and was then sustained. VSMC proliferation after balloon catheter injury of the rat iliac artery is associated with coordinated expression of positive (cdk2, cyclin E and PCNA) and negative (p21, p27) regulators. Cell cycle regulators such as cdk2, cyclin E, p21, p27 may be suitable targets for the control of intimal hyperplasia.     

Platelet-complement interactions in mesangial proliferative nephritis in the rat.

Complement has been reported to mediate mesangiolysis and glomerular hypercellularity in the rat in a model of glomerulonephritis (GN) induced with anti-Thy 1 antibody. To investigate the mechanism for the complement-mediated hypercellularity, the authors first determined if the effect of complement depletion was to inhibit cell proliferation or whether the effect was primarily to inhibit leukocyte infiltration. Rats depleted of complement with cobra venom factor (CVF) had 1) significantly less mesangiolysis than controls at day 5 (0.6 +/- 0.1 versus 3.4 +/- 0.4, scale 0-4+, P less than 0.001); 2) less cell proliferation, as assessed by immunostaining for the proliferating cell nuclear antigen (PCNA)/cyclin, a cell-cycle-dependent antigen (0.5 +/- 0.1 versus 2.4 +/- 0.7 cells/glomerular cross-section, P less than 0.01); and 3) less leukocyte infiltration as assessed by immunohistochemical labeling (0.6 +/- 0.1 versus 1.9 +/- 0.3 cells/glomerular cross-section, P less than 0.01). Because it was reported recently that platelets also mediate glomerular cell proliferation in this model, this study examined whether the mechanism for complement-mediated cell proliferation involved an effect on glomerular platelet localization. The glomerular uptake of 111In-labeled platelets was quantitated in normal and CVF-treated rats at 1, 4, 12, and 24 hours after induction of GN. Rats with anti-Thy 1 GN had substantial glomerular accumulation of platelets at all times studied, peaking at 4 hours (608 +/- 171 platelets per glomerulus). Complement depletion profoundly reduced glomerular platelet localization in anti-Thy 1 GN (mean less than 35 platelets per glomerulus at all times studied, P less than 0.05). Thus these studies demonstrate an important role for complement in mediating platelet localization in anti-Thy 1 GN, an effect that may account for the complement-dependent, neutrophil-independent glomerular hypercellularity in this model.     

The rapamycin derivative RAD inhibits mesangial cell migration through the CDK-inhibitor p27KIP1

The link between mesangial cell (MC) proliferation and migration during glomerular repair in the experimental mesangial proliferative glomerulonephritis suggests that cell cycle regulation and cell migration require similar pathways, such as cell cycle proteins. The immunosuppressant RAD inhibits mesangial cell (MC) proliferation via G1/S arrest. Moreover, RAD dramatically impairs glomerular healing in the anti-Thy1 model. We tested the hypothesis that RAD alters MC migration in vitro and that this effect was mediated by the CDK-inhibitors p21(CIP1) and p27(KIP1). Using a modified Boyden chamber in vitro migration assay, our results showed that RAD dose dependently (1-50 nM) inhibited fibronectin-induced chemotaxis in wild-type (wt) MC. RAD treatment prevented the decrease in p27(KIP1) induced by mitogenic growth factors, but had no effect on p21(CIP1) by Western blot analysis. The antimigratory effect of RAD in wt MC was substantially dependent on p27(KIP1), but not p21(CIP1), since the inhibitory effects of 1-10 nM RAD on MC migration were similar in p21(CIP1) deficient and wild-type MC. The effect of RAD on MC migration was also examined in the anti-Thy1 model by BrdU-labeling of proliferating MC on day 3 that typically repopulate the glomerulus from the hilus. A control biopsy on day 3 was taken to define the starting point prior to the initiation of RAD (3 mg/kg or placebo). MC migration was determined on day 7 by measuring the distances of BrdU-labeled MC (OX-7+/BrdU+cells) from the glomerular hilus using computerized morphometry. RAD significantly reduced the migratory response of BrdU-labeled MC compared to controls.We conclude that the immunosuppressant RAD effectively inhibits MC migration in vivo and in vitro thereby limiting the normal glomerular repair process after severe injury. Moreover, RAD-induced inhibition of MC migration in vitro is partially mediated by the CDK-inhibitor p27(KIP1), but not p21(CIP1).     

Interleukin-10 inhibits experimental mesangial proliferative glomerulonephritis

Conflicting reports exist regarding the effects of interleukin-10 (IL-10) on mesangial cells. There have been reports of both proliferative and antiproliferative effects, and both proinflammatory and anti-inflammatory effects of IL-10 on mesangial cells. However, the potential for IL-10 to affect glomerulonephritis characterized by mesangial proliferation is not known. To test the hypothesis that IL-10 would limit experimental mesangial proliferative glomerulonephritis, IL-10 was administered to rats in which mesangial proliferative glomerulonephritis was induced by administration of anti-Thy 1 antibody. Compared to control treated rats, IL-10 treated rats showed less proliferation, with fewer cells in glomeruli. Glomerular cellular proliferation was reduced, assessed by the numbers of cells within glomeruli expressing either proliferating cell nuclear antigen (PCNA) or bromodeoxyuridine. Glomerular macrophage influx (but not the proportion of glomerular macrophages that were PCNA positive) was reduced by IL-10 administration. There was no significant reduction in glomerular alpha-smooth muscle actin staining. IL-10 treatment resulted in reduced renal IL-1beta mRNA expression and reduced glomerular ICAM-1 expression, but renal expression of MCP-1 and osteopontin mRNA was unaltered. This study demonstrates that in experimental mesangial proliferative glomerulonephritis IL-10 diminishes inflammatory cell recruitment and mesangial cell proliferation. The effects of IL-10 in inhibiting mesangial cell proliferation are likely to be due to a combination of direct effects of IL-10 on mesangial cells and effects mediated by macrophages.     

Akt／NF-kB信号途径在免疫复合物诱导肾系膜细胞增殖中的作用

目的：观察免疫复合物(IC)能否诱导体外培养的肾小球系膜细胞(MC)增殖及其Akt/NF—kB信号在其中的作用。方法：实验设对照组、刺激组及Akt1正义(SODN)、错义(MSODN)与反义寡核苷酸(ASODN)组。寡核苷酸组：以lipofectin分别介导Akt1 SODN、MSODN、ASODN转染MC8h；对照组和刺激组：以lipofectin作用MC8h。然后刺激组和寡核苷酸组均用聚合IgG(aggregated IgG，AIgG，一种标准的IC模型)刺激，对照组用等量的单体IgG刺激。用MTT比色法检测细胞的增殖，流式细胞术分析MC的细胞周期，RT-PCR分析mRNA的表达，Westem blot检测蛋白的表达，电泳迁移率变动实验(EMSA)检测NF—KB的活性。结果：IC可诱导MC中NF-kB的活化，上调细胞周期蛋白cyclinD1的mRNA及蛋白表达，促进MC进入S期。Akt1 ASODN可抑制Akt1蛋白的表达，抑制IC诱导的MC中NF—kB活性，抑制cyclinD1 mRNA及其蛋白的表达，进而抑制IC促进MC进入S期和增殖。Akt1 SODN、MSODN则无上述抑制作用。结论：IC可通过Akt/NF—kB信号途径，介导体外培养的MC增殖。提示Akt／NF-kB信号途径，可作为干预IC诱导MC过度增生的一个新的治疗靶点。     

SC58125对HepG-2细胞增殖和凋亡的影响

目的：探讨SC-58125对HepG-2细胞增殖和凋亡的作用及其分子机理。方法：应用细胞培养、MTT、TUNEL、流式细胞光度术、琼脂糖凝胶电泳及Western blot等方法研究SC-58125对HepG-2细胞增殖和凋亡的作用及其分子机理。结果：SC58125抑制HepG-2细胞的增殖、诱导其凋亡及引起G₀/G₁期阻滞，S期抑制。并使P33^cdk2、P34^cdc2、cyclinB₁、cyclinE、Mpm-2、Rb、PCNA 7种蛋白水平下降。结论：SC58125抑制HepG-2细胞的增殖及诱导其凋亡，可能与P33^cdk2、P34^cdc2、cyclinB₁、cyclinE、Mpm-2、Rb、PCNA 7种蛋白水平的下降有关。     

细胞周期蛋白依赖性激酶cdk2和cdk4在前列腺增生和癌变组织中的表达

目的：探讨细胞周期蛋白依赖性激酶ｃｄｋ２和ｃｄｋ４在前列腺增生（ＢＰＨ）和前列腺癌（ＰＣ）的发生发展过程中作用及其与ＰＣＮＡ之间关系。方法：应用免疫组化ＳＰ法检测１８例正常前列腺（ＮＰ）、６２例ＢＰＨ和３３例ＰＣ组织中ｃｄｋ２、ｃｄｋ４和ＰＣＮＡ的表达。结果：前列腺上皮和间质组织中均见ｃｄｋ２和ｃｄｋ４表达。ＮＰ中两者表达均分别显著低于ＢＰＨ和ＰＣ;ＢＰＨ的上皮细胞中两者表达均分别显著低于ＰＣ,但ＢＰＨ的间质细胞中两者表达与ＰＣ相比均无显著性差异。ＢＰＨ和ＰＣ中ｃｄｋ２及ｃｄｋ４表达与ＰＣＮＡ指数均呈正相关。结论：ｃｄｋ２和ｃｄｋ４异常表达参与ＢＰＨ和ＰＣ的发生发展过程,其可能是通过改变细胞周期及促进细胞异常增殖而起作用的。     

Relationship between the expression of cyclins/cyclin-dependent kinases and sex-steroid receptors/Ki67 in normal human endometrial glands and stroma during the menstrual cycle

Cell cycle regulatory molecules were analysed in normal humanendometrial tissue using antibodies against cyclins D1, E, A,and B1 and cyclin-dependent kinases (CDKs) cdk4, cdk2, and cdc2.The expression of these regulatory molecules in gland cellsand stromal cells was compared with the expression of oestrogenreceptors (ER), progesterone receptors (PR), and Ki67 (a growth-relatedmolecule). In general, a substantially higher percentage ofthe gland cells stained positive for cyclins and CDKs duringthe proliferative phase of the menstrual cycle. Cyclin E, cdk2and/or cdk4 were especially apparent in the cytoplasm of mostof the gland cells as well as in the stromal cells. In contrast,most of the regulatory molecules were undetectable in the glandcells by the end of the secretory phase of the cycle, but theydid not decline in the stromal cells. The data also revealedthat ER, PR, and Ki67 in both gland cells and stromal cellsfollow the same basic pattern of expression as the cyclins andCDKs. These results suggest that cyclins and CDKs are functionallyinvolved in the rhythmic proliferation of normal human endometrialtissue, and the action of these agents may be related to theendometrial levels of sex steroids and Ki67. cyclin/cyclin-dependent kinase/endometrium/Ki67/steroid receptors     

E2F陷阱DNA对前列腺癌PC-3M细胞凋亡的诱导

目的：为观察转录因子E2F陷阱DNA 对雄激素非依赖性前列腺癌细胞PC-3M增殖和凋亡的影响。方法：采用脂质体lipofectamine将E2F decoy DNA、ARE decoy DNA和control decoy DNA分别转染PC-3M细胞，MTT检测其对细胞增生的影响，倒置相差显微镜观察细胞形态变化，流式细胞术检测细胞凋亡率，并进行染色体DNA断裂的测定；通过RT-PCR检测转染的PC-3M细胞中c-Myc mRNA、cyclin D1 mRNA表达水平的变化，通过Western blotting检测细胞中c-Myc蛋白、cyclin D1蛋白表达水平的变化。结果：E2F decoy DNA转染后的PC-3M细胞的生长受到明显抑制;转染后的细胞形态变化符合凋亡的典型改变，染色体断裂明显,细胞凋亡率为26.35%；c-Myc、cyclin D1的表达受到抑制。结论：E2F decoy DNA可诱导雄激素非依赖性前列腺癌PC-3M凋亡和抑制细胞增殖,其机制可能涉及到c-Myc mRNA、cyclin D1的表达变化。     

Cell cycle differences in vitro between primitive hematopoietic cell populations from adult and umbilical cord blood

Lineage-negative (Lin(-)) cell populations, obtained by negative selection from umbilical cord blood (UCB) and adult mobilized peripheral blood (aMPB), were cultured in serum-free liquid cultures supplemented with a mixture of seven stimulatory cytokines. On specific days, proliferation potential was assessed and cell cycle status was determined by DNA content. Expression of the cell cycle regulators cyclin D3 (cD3), cyclin-dependent kinase 4 (cdk4), p21(cip1/waf1) (p21), and p27(kip1) (p27) was also determined. As expected, UCB cells showed significantly higher proliferation potentials than aMPB cells, particularly during the first 7 days of culture. During this period of time, higher numbers of cell cycles were observed in UCB cells (7-9 cycles), as compared to aMPB cells (5-6 cycles). Higher levels of cD3, cdk4, and p27 were also detected in UCB cells. Our results confirm that UCB cells possess an intrinsically higher proliferation potential, as compared to aMPB cells, and suggest that such a biological difference is due, at least in part, to differences in cell cycle status. This, in turn, seems to result from the differential expression of cell cycle regulatory molecules.     

Transgelin is a marker of repopulating mesangial cells after injury and promotes their proliferation and migration

Mesangial cell (MC) migration is essential during glomerular repair and kidney development. The aim of the study was to identify marker/player for glomerular progenitor/reserve cells migrating into the glomerulus after MC injury and during glomerulogenesis in the rat. Experimental mesangial proliferative nephritis was induced in Sprague Dawley rats by intravenous injection of OX-7 antibody. We investigated mRNA expression profiles in isolated glomeruli from on days 0, 1, 2, 3, and 5 after induction of anti-Thy1 nephritis using Affymetrix microarray technology. Using self-organizing maps, transgelin was identified as a new marker for repopulating glomerular cells. Expression of transgelin during anti-Thy1 nephritis was investigated by northern blot, real-time PCR, western blot, and immunohistochemistry. Migration and proliferation assays using isolated MCs after transgelin knockdown by siRNA were performed to investigate the potential role of transgelin during glomerular repopulation. Transgelin mRNA was not detected in healthy glomeruli. It was strongly upregulated during the repopulation process starting on day 1, continued to be increased until day 5 and disappeared on day 7. Transgelin was specifically expressed at the edge of the migratory front during glomerular repopulation as indicated by transgelin/OX-7 double staining. Transgelin expression was similar in migrating vs non-migrating MCs in vitro. Blocking of transgelin expression by siRNA treatment resulted in inhibition of MC migration and proliferation. Transgelin was also expressed in MCs during glomerulogenesis and in biopsies from patients with IgA nephritis. In conclusion, transgelin in the kidney is upregulated in repopulating MCs in vivo and supports their migratory and proliferative repair response after injury.     

Suppressor activity of anergic T cells induced by IL-10-treated human dendritic cells: association with IL-2- and CTLA-4-dependent G1 arrest of the cell cycle regulated by p27Kip1

We have previously shown that human IL-10-treated dendritic cells (DC) induce an antigen-specific anergy in CD4+ T lymphocytes. These anergic T cells are characterized by an inhibited proliferation, a reduced production of IL-2, and additionally display antigen-specific suppressor activity. In this study we investigated the mechanisms underlying the anergic state and regulatory function of these T cells. We did not observe enhanced rates of programmed cell death of anergic CD4+ suppressor T cells compared to T cells stimulated with mature DC. Cell cycle analysis by DNA staining and Western blot experiments revealed an arrest of anergic CD4+ T suppressor cells in the G1 phase. High levels of the IL-2-dependent cyclin-dependent kinase (cdk) inhibitor p27Kip1 were found in anergic CD4+ suppressor T cells resulting in an inhibited activation of retinoblastoma protein and an arrest of cell cycle progression in the G1 phase. Addition of IL-2, but not blocking of the CTLA-4 pathway restored the proliferation of the suppressor T cells. In contrast, both treatments induced a down-regulation of p27Kip1 and acomplete inhibition of the antigen-specific regulatory function as demonstrated by high proliferation and enhanced IFN-gamma production of co-cultured T cells. Further experiments demonstrated that p27Kip-expressing regulatory CD4+CD25+ T cells did not contribute to induction of T cell anergy in this model. Our data show that regulatory function of anergic CD4+ suppressor T cells is associated with an arrest in the G1 phase of the cell cycle mediated by increased levels of the IL-2- and CTLA-4-dependent cdk inhibitor p27Kip1.     

Molecular basis for the lack of T cell proliferation induced by an altered peptide ligand

In this report, we explore the mechanisms underlying cell cycle progression in T cells stimulated with an altered peptide ligand (APL) versus wild-type peptide. APL stimulation did not induce proliferation compared to wild-type peptide stimulation. To determine the point at which cell cycle progression is blocked, we have examined molecules responsible for regulating the retinoblastoma tumor suppressor gene product, pRb, which in its active state prevents G1/S progression. The majority of cells stimulated with an APL did not progress beyond G1; however, a small population did make the G1/S transition. These few cells passed the late G1 restriction point, divided and subsequently arrested at the next G1 phase. The lack of sustained signaling events following stimulation with an APL failed to induce cyclin E:cdk2 activity, a regulator which hyper-phosphorylates and inactivates pRb. Exogenous IL-2 addition did not compensate for the lack of proliferation following APL stimulation. Furthermore, the inability of the cells to enter S phase during partial T cell activation cannot be accounted for by p27Kip1 inhibition of cyclin E:cdk2 complexes. Upon APL stimulation, an increase in association of p27Kip1 with cyclin E:cdk2 complex was not observed, suggesting that instead, decreased cyclin E:cdk complex formation might contribute to the failure to progress from G1/S. Therefore, while for a majority of cells, wild-type stimulation results in cell cycle progression, APL stimulation is not sufficient to drive cells beyond G1.      

G1 cyclin-dependent kinases are insufficient to reverse dE2F2-mediated repression

Here we show that the cell cycle defects of dE2F1-depleted cells depend on the cooperative effects of dE2F2 and DACAPO (DAP), an inhibitor of Cyclin E/cyclin-dependent kinase 2 (CycE/cdk2). The different properties of cells lacking dE2F1/dE2F2 and dE2F1/DAP lead to the surprising observation that dE2F2-mediated repression differs from retinoblastoma family protein 1 (RBF1) inhibition of dE2F1, and is resistant to both CycE/cdk2 and Cyclin D/cyclin-dependent kinase 4 (CycD/cdk4). This resistance occurs even though dE2F2/RBF1 complexes are disrupted by CycE/cdk2, and may explain why dE2F2 is so potent in the absence of de2f1. The implication of these results is that cells containing dE2F2 require dE2F1 to either prevent, or reverse, dE2F-mediated repression.     

Distributional changes of BrdU, PCNA, E2F1 and PAL31 molecules in developing murine palatal rugae

The distribution of cells incorporating bromodeoxyuridine (BrdU) and the expression of molecules involved in the control of cell proliferation (proliferating cell nuclear antigen [PCNA], a cellular factor in F9 teratocarcinoma cells that recognizes an adenovirus E1A inducible promoter 1 [E2F1] and proliferation-related acidic nuclear protein 31 [PAL31]) during morphogenesis of the murine palatine rugae (PR) was examined histochemically. Pattern formation of the PR rudiment was initiated with cell cycle related molecules in the epithelium of the primary palate. Cells which had incorporated BrdU were detected at the outer areas of the presumptive epithelial placode (EP) and the EP at 11.5–13.5 days post coitum (dpc) and the outer areas of the PR protrusion after 14.5 dpc. The number of PCNA-positive cells at the central area of the PR protrusion decreased after 16.5 dpc. E2F-positive cells were detected at the outer areas of the PR protrusion at 15.5 and 16.5 dpc. The number of PAL31-positive cells at the presumptive EP area and the already-formed EP area was decreased at 11.5–13.5 dpc. In two dimensional histological reconstructions, PAL31 expression approximately corresponded to the distribution of BrdU-positive cells at 11.5 and 13.5 dpc. EP placode formation might be regulated by spatiotemporal cell proliferation control involving the expression of the PAL31 molecule. Following EP formation, PR development and growth control involved the expression of E2F1 and PCNA molecules.