Interrelationships between strains of Salmonella enteritidis belonging to phage types 4, 7, 7a, 8, 13, 13a, 23, 24 and 30

Salmonella enteritidis strain P278849 expressed long-chain lipopolysaccharide (LPS, termed 'smooth'), carried plasmids of 38, 34 (pDEP 44, incompatibility group N, R-type AS), 2.0 and 1 MDa, and belonged to phage type (PT) 23. Introduction of pDEP 44 into a smooth strain of Salm. enteritidis PT 4 produced a smooth strain of Salm. enteritidis of PT 24. Transfer of this plasmid into a strain of PT 8 also resulted in formation of a smooth strain of Salm. enteritidis of PT 24. Moving pDEP 44 into strains of Salm. enteritidis of PTs 7 or 7a which did not express LPS (termed 'rough') resulted in rough strains of PT 23. In contrast, transfer of this plasmid into a smooth strain of Salm. enteritidis PT 7a resulted in a smooth strain of PT 23. Introduction of pDEP 44 into strains of Salm. enteritidis of PT 13 or PT 13a did not change the phage type, whereas transferring the plasmid into strains of PT 30 caused strains to become resistant to lysis by the typing phages and therefore untypable. The possibility of strains of Salm. enteritidis of PT 8 being the progenitors of strains of Salm. enteritidis of PT 24 must now be considered when investigating the epidemiology of Salm. enteritidis of PT 24 infections in areas where Salm. enteritidis PT 8 is common.     

Salmonella enterica serotype Enteritidis phage types 4, 7, 6, 8, 13a, 29 and 34: a comparative analysis of genomic fingerprints from geographically distant isolates

AIMS: To evaluate genetic heterogeneity in the most common phage types of Salmonella enterica serovar Enteritidis. METHODS AND RESULTS: A total of 233 isolates of Salm. Enteritidis from England, Northern Ireland, Spain, Hong Kong and the USA belonging to phage types (PT) 4 (n=88), PT7 (n=12), PT6 (n=72), PT8 (n=14), PT13a (n=29), PT29 (n=14) and PT34 (n=4) were characterized by PstI-SphI (PS) ribotyping and pulsed-field gel electrophoresis after digestion of DNA with XbaI. PS ribotyping differentiated the isolates into 53 different PS types and PFGE showed 14 different macrorestriction profiles; with the combination of both methods, 73 combined types were identified. Some of these clones appeared to be present within several countries. Movement of foodstuffs, animals or people may have been involved in the spread of these strains. On the other hand, some clones were only found in specific locations. CONCLUSIONS: Several well defined clonal lines seem to co-exist within the different phage types included in this study, and a combined typing approach may constitute a useful tool for epidemiological investigations. Clustering analysis of ribotypes and PFGE types agree with previous studies and suggest that phage types that share receptor binding properties can be distinguished as two families: the PT4 family including PT7 and PT6, and the PT8 family including PT13a. The other phage types are difficult to place in a family unless the geographical site of isolation is known. SIGNIFICANCE AND IMPACT OF THE STUDY: This paper reports on an extensive assessment of the use of molecular tools for the study of the epidemiology of the enteric pathogen Salm. Enteritidis. It also gives new information regarding relationships among some common phage types.     

Classification,Prediction, and Verification of the Regioselectivity of Fungal Polyketide Synthase Product Template Domains

The fungal iterative nonreducing polyketide synthases (NRPKSs) synthesize aromatic polyketides, many of which have important biological activities. The product template domains (PT) embedded in the multidomain NRPKSs mediate the regioselective cyclization of the highly reactive polyketide backbones and dictate the final structures of the products. Understanding the sequence-activity relationships of different PT domains is therefore an important step toward the prediction of polyketide structures from NRPKS sequences and can enable the genome mining of hundreds of cryptic NRPKSs uncovered via genome sequencing. In this work, we first performed phylogenetic analysis of PT domains from NRPKSs of known functions and showed that the PT domains can be classified into five groups, with each group corresponding to a unique product size or cyclization regioselectivity. Group V contains the formerly unverified PT domains that were identified as C6-C11 aldol cyclases. The regioselectivity of PTs from this group were verified by product-based assays using the PT domain excised from the asperthecin AptA NRPKS. When combined with dissociated PKS4 minimal PKS, or replaced the endogenous PKS4 C2-C7 PT domain in a hybrid NRPKS, AptA-PT directed the C6-C11 cyclization of the nonaketide backbone to yield a tetracyclic pyranoanthraquinone 4. Extensive NMR analysis verified that the backbone of 4 was indeed cyclized with the expected regioselectivity. The PT phylogenetic analysis was then expanded to include ∼100 PT sequences from unverified NRPKSs. Using the assays developed for AptA-PT, the regioselectivities of additional PT domains were investigated and matched to those predicted by the phylogenetic classifications.     

Cell organization of the rat pars tuberalis. Evidence for open communication between pars tuberalis cells, cerebrospinal fluid and tanycytes

The pars tuberalis (PT) is the only pituitary region in close contact with the medial-basal hypothalamus and bathed by cerebrospinal fluid (CSF). Although PT has long been recognized as an endocrine gland, certain aspects of its structure remain obscure. The present investigation has been designed to gain information concerning (1) the cellular organization of PT, (2) the PT/median eminence spatial relationship and (3) the exposure of various cell compartments of PT to CSF. Non-endocrine cells (S100-reactive) appear as the organizer of the PT architecture. The apical poles of these cells line large cistern-like cavities and the processes of these cells establish a close spatial relationship with PT-specific secretory cells, portal capillaries and tanycytes. The cisterns are also endowed with clusters of ciliated cells and with a highly electron-dense and PAS-reactive content. The unique spatial organization of endocrine and non-endocrine cells of the PT supports a functional relationship between both cell populations. PT endocrine cells display a hallmark of PT-specific cells, namely, the paranuclear spot, which is a complex structure involving the Golgi apparatus, a large pool of immature secretory granules and a centriole from which originates a single 9+0 cilium projecting to the intercellular channels. Horseradish peroxidase (HRP) injected into the CSF readily reaches the intercellular channels of PT and the inner channel of the single cilium and is incorporated by the endocytic machinery of the secretory cells. The PT endocrine cells, through their single 9+0 cilium, may act as sensors of the CSF. HRP also reaches the lumen of the cisterns, indicating that this PT compartment is also exposed to CSF. PT endocrine cells establish direct cell-to-cell contacts with hypothalamic β₁ tanycytes, suggesting a second means of brain-PT communication.     

Pterostilbene,a natural small-molecular compound,promotes cytoprotective macroautophagy in vascular endothelial cells

Chemical modulators of macroautophagy (herein referred to as autophagy) have aroused widespread interest among biologists and clinical physicians because of their potential for disease therapy. Pterostilbene (PT), a natural small-molecular compound, has been demonstrated to inhibit oxidized low-density lipoprotein (oxLDL)-induced apoptosis in vascular endothelial cells (VECs). The aim of the present study was to investigate whether and how PT could induce VEC autophagy. PT at 0.5 or 1 μM could effectively induce autophagosome formation in human umbilical vein VECs (HUVECs). PT promoted autophagy via a rapid elevation in intracellular calcium ([Ca^2 +]_i) concentration and subsequent AMP-activated protein kinase α1 subunit (AMPKα1) activation, which in turn inhibited mammalian target of rapamycin, a potent inhibitor of autophagy. PT-induced AMPKα1 activation and autophagy were refractory to the depletion of serine/threonine kinase 11 but depended on calcium/calmodulin-dependent protein kinase kinase-β activation. Interestingly, PT stimulated cytoprotective autophagy so as to aid in the removal of accumulated toxic oxLDL and inhibit apoptosis in HUVECs. Our study provides a potent small molecule enhancer of autophagy and a novel useful tool in exploring the molecular mechanisms for crosstalk between apoptosis and autophagy. PT could serve as a potential lead compound for developing a class of autophagy regulator as autophagy-related diseases therapy.     

Prothrombin binds to the surface of apoptotic,but not viable,cells and serves as a target of lupus anticoagulant autoantibodies

Anti-phospholipid Ab (aPL) are a heterogeneous group of autoantibodies directed against various combinations of phospholipids (PL) and PL-binding proteins. Lupus anticoagulant (LA) Ab, a subset of aPL, exhibit anticoagulant properties in vitro, but are procoagulant in vivo. Most LA Ab are specific for either beta(2)-glycoprotein I (beta(2)GPI) or prothrombin (PT), two PL-binding proteins. We have previously shown that beta(2)GPI and beta(2)GPI-dependent aPL bind specifically to apoptotic, but not viable, thymocytes. In this study, we demonstrate that PT, like beta(2)GPI, binds selectively to the surface of apoptotic, but not viable, Jurkat cells. Furthermore, PT supports the binding of systemic lupus erythematosus-derived polyclonal and murine monoclonal LA Ab to apoptotic cells. Two LA mAb, which differed dramatically in their relative affinities for PT, were studied. Although one mAb (29J3-62) had a high affinity for PT alone, the other (29I4-24) showed minimal reactivity with PT alone and required PL for elevated binding. Monovalent fragments of 29I4-24 reacted with PL-bound PT with high affinity, suggesting that this mAb recognizes a PL-dependent epitope. Despite these differences, PT-dependent binding of both mAb to apoptotic cells was 30-fold greater than that to viable cells. Moreover, binding of PT to apoptotic cells was, itself, increased in the presence of bivalent, but not monovalent, forms of either mAb. In summary, our data demonstrate the following: 1) specific binding of PT to apoptotic cells, an effect enhanced by PT-dependent LA Ab; 2) heterogeneity of PT-dependent LA Ab; and 3) potential pathogenicity of Ab of either low or high affinity for PT.     

Interaction with gravitropism,reversibility and lateral movements of phototropically stimulated potato shoots

Phototropic (PT) and gravitropic (GT) bending are the two major tropic movements that determine the spatial position of potato shoots. We studied PT bending of potato plantlets grown under long-day photoperiods in several prearranged position setups providing different interactions with the GT response. Starting with the standard PT stimulation setup composed of unilateral irradiation of vertically positioned shoots, experiments were also done in antagonistic and synergistic setups and in treatments with horizontal displacement of the light source. In the standard setup, PT bending suppressed the GT bending, which could occur only if the PT stimulation was cancelled. The antagonistic position, with phototropism and gravitropism attempting to bend shoots in opposite directions, showed phototropism and gravitropism as independent bending events with the outcome varying throughout the day reflecting diurnal changes in the competence of individual tropic components. Whilst gravitropism was constant, phototropism had a marked daily fluctuation of its magnitude with a prominent morning maximum starting an hour after the dawn in the growth room and lasting for the next 6 h. When phototropism and gravitropism were aligned in a synergistic position, stimulating shoot bending in the same direction, there was little quantitative addition of their individual effects. The long period of morning PT bending maximum enabled multiple PT bending events to be conducted in succession, each one preceded by a separate lag phase. Studies of secondary PT events showed that potato plantlets can follow and adjust their shoot position in response to both vertical and horizontal movements of a light source. PT bending was reversible, since the 180° horizontal change of a blue light (BL) source position resulted in reversal of bending direction after a 20-min-long lag phase.     

The effects of plyometric, tennis-drills, and combined training on reaction, lateral and linear speed, power, and strength in novice tennis players

Reaction time, first-step quickness, lateral (side steps), and forward speed over short distances are important parameters for tennis performance. The aims of this study were: (i) to diagnose the presence of laterality in tennis lateral movements and (ii) to compare the effects of plyometric training (PT), tennis-specific drills training (TDT), and combined training (CT) on performance in tennis-specific movements and power/strength of lower limbs. Sixty-four novice tennis players (21.1 +/- 1.3 years) were equally (n = 16) assigned to a control (C), PT, TDT, or CT. Training was performed 3 times/week for 9 weeks. Testing was conducted before and after training for the evaluation of reaction time (single lateral step), 4-m lateral and forward sprints, 12-m forward sprints with and without turn, reactive ability, power, and strength. There was a significant difference in lateral speed (side-steps) between the 2 sides (P < 0.05). PT, TDT, or CT improved the 4m lateral and forward sprints (P < 0.05). PT and CT improved also the reaction time of the "slow" side (P < 0.05), whereas TDT and CT improved the 12-m sprint performances with and without turn (P < 0.05). Power and strength improved in most tests after PT and CT. Lateral and forward sprints were correlated (r = -0.50 to -0.75; P < 0.05) with power/strength. In conclusion, PT improved fitness characteristics that rely more on reactive strength and powerful push-off of legs such as, lateral reaction time, 4-m lateral and forward sprints, drop jump and maximal force. TDT improved all 4-m and 12-m sprint performances, whereas CT appeared to incorporate the advantage of both programs and improved most tests items. Tennis coaches should be aware that each training regimen may induce more favorable changes to different aspects of fitness.     

The Five-Point Test: Reliability,Validity and Normative Data for Children and Adults

The present study provides normative data from a sample of 257 healthy children and 608 adults on a modified version of the Five-Point Test (5PT). The 5PT is a structured and standardized test measuring figural fluency functions. Interrater reliability, test-retest-reliability and construct validity of this measure were analyzed. The sensitivity of the task for cognitive disturbances of patients with neurological diseases was proven by analyzing the test performance in the 5PT of patients with Parkinson''s disease. Finally, normative data stratified by age and corrected for education level is provided. The results of the present study confirm the value of the 5PT in the measurement of figural fluency functions in clinical examination and neuropsychological research.     

Tumor cell targeting of transferrin-PEG-TNF-alpha conjugate via a receptor-mediated delivery system: design, synthesis, and biological evaluation

PEGylation is a procedure of growing interest for enhancing the therapeutic and biotechnological potential of peptides and proteins. Transferrin (Tf) has been proposed to be useful for targeting cancer cells. The aim of this study was to modify PEGylated recombinant human tumor necrosis factor alpha (PEG-TNF-alpha) with Tf to form Tf-PEG-TNF-alpha conjugates, which would maintain the advantages of PEGylation and also achieve the function of active targeting to tumor cells. In PEGylation reactions with 5-, 20-, 40-, and 60-fold molar excess of 3.4 kDa N-hydroxysuccinimide-PEG-maleimide (PT1, PT2, PT3, and PT4, respectively), PEG-TNF-alpha conjugates with different PEG chains were synthesized. A perfusion chromatography technique using a cation-exchange column was introduced to purify PEG-TNF-alpha conjugates. PT4 with about five PEG chains was selected as a lead candidate due to highest extent of PEGylation and maximum reaction yield. Thiolated Tf was conjugated to the maleimide group at the distal end of the PEG chains on the PEG-TNF-alpha conjugates, with the resulting Tf-PEG-TNF-alpha conjugates after purification containing approximately one Tf ligand on one TNF-alpha molecule. The conjugate of Tf and PT4 (TPT4) was selected to assess the specificity and affinity to transferrin receptor (TfR) on two kinds of tumor cells, K562 and KB. Both the receptor binding assays and the competition experiments were performed using radioligand binding analysis. The results demonstrated that TPT4 as well as Tf bound specifically to the TfR on the tumor cell surface and the affinity of the conjugate to TfR was similar to that of native Tf. In contrast, PEG-TNF-alpha demonstrated no specificity. The biodistribution and antitumor effects were investigated in S-180 tumor-bearing mice. It was found that TPT4 could markedly alter in vivo behavioral characteristics of TNF-alpha. Compared with TNF-alpha and PT4, extravasated TPT4 in tumor tissues exhibited a significantly delayed blood clearance and the highest intratumoral TNF-alpha levels. Furthermore, the inhibitory rate of tumor of TPT4 enhanced 5.3- and 1.8-fold over that of TNF-alpha and PT4, indicating that TPT4 exhibited the highest antitumor activity. These results suggested that Tf-PEG-TNF-alpha was a useful long circulating conjugate with the capabilities of specific receptor binding resulting in enhanced antitumor activity of TNF-alpha.     

Glucocorticoid receptor-induced down-regulation of MMP-9 by ginseng components, PD and PT contributes to inhibition of the invasive capacity of HT1080 human fibrosarcoma cells

We examined the effects of the purified ginseng components, panaxadiol (PD) and panaxatriol (PT), on the expression of matrix metalloproteinase-9 (MMP-9) in highly metastatic HT1080 human fibrosarcoma cell line. A significant down-regulation of MMP-9 by PD and PT was detected by Northern blot analysis. However, the expression of MMP-2 was not changed by treatment with PD and PT. Quantitative gelatin based zymography confirmed a markedly reduced expression of MMP-9, but not MMP-2 in the treatment of PD and PT. To investigate whether the reduced level of MMP-9 by PD and PT affects the invasive capacity of HT1080 cells, we conducted an in vitro invasion assay with PD and PT treated cells. The results of the in vitro invasion assay revealed that PD and PT reduced tumor cell invasion through a reconstituted basement membrane in the transwell chamber. Because of the similarity of chemical structure between PD, PT and dexamethasone (Dexa), a synthetic glucocorticoid, we investigated whether the down-regulation of MMP-9 by PD and PT were mediated by the nuclear translocation of glucocorticoid receptor (GR). Increased GR in the nucleus of HT1080 human fibrosarcoma cells treated by PD and PT was detected by immunocytochemistry. Western blot and gel retardation assays confirmed the increase of GR in the nucleus after treatment with PD and PT. These results suggest that GR-induced down-regulation of MMP-9 by PD and PT contributes to reduce the invasive capacity of HT1080 cells.     

Phyllodes tumour: cytologic and histologic presentation of 22 cases, and immunohistochemical demonstration of p53

A retrospective study of 22 cases of phyllodes tumour (PT) was undertaken to evaluate the potential value of fine needle aspiration (FNA) cytology in the diagnosis of benign and borderline PT. Histological material was available from 12 patients with typical benign PT (group 1), six patients with less typical changes (group 2) and four cases of borderline PT (group 3). Cytological presentation of PT in these cases was similar to that described by other cytologists, although abundant cellular material was obtained in only eight FNAs, naked nuclei were present in nine cases only, and atypical or suspicious cytological features were found in seven cases. Comparative analysis of p53 was made in nine patients with PT, five cases with other benign breast lesions and five with malignant lesions. p53 reaction was positive in five of nine patients with PT (all cases from groups 2 and 3), compared with two of five cases of carcinoma. p53 was negative in all patients with PT from group 1 and the five other benign cases. We suggest that cytopathologists should be careful when a myxoid stromal component is present in cytological smears.     

Pulmonary trunk, ductus arteriosus, and pulmonary arterial phasic blood flow interactions during systole and diastole in the fetus

Although the distribution of average fetal pulmonary trunk (PT) blood flow favors the ductus arteriosus (DA) over the lungs, the phasic aspects of this distribution during systole and diastole are not well understood. Accordingly, flow profile and wave intensity (WI) analyses were performed at baseline and during brief flow increases accompanying an extrasystole (ES) in 10 anesthetized late-gestation fetal sheep instrumented with PT, DA, and left pulmonary artery (PA) micromanometer catheters and transit-time flow probes. At baseline, 83% of mean PT flow crossed the DA and 17% entered the lungs. However, early systolic flow associated with a forward-running compression wave (FCW(is)) was higher in the PA and predominant DA flow only emerged in midsystole when a large PA backward-running compression wave (BCW(ms)), which reduced PA flow, was transmitted into the DA as a forward-running compression wave (FCW(ms)) that increased flow. Subsequent protodiastolic forward DA flow occurring during pulmonary valve closure was associated with substantial retrograde PA flow, but insignificant PT flow. Conversely, forward DA flow in the remainder of diastole occurred with forward PT but near-zero PA flow. These flow and WI patterns, in conjunction with the results of mathematical modeling, suggest that 1) fetal PT flow preferentially passes into the PA during early systole due to a lower PA-than-DA characteristic impedance, while DA flow predominates in mid- and late systole due to flow effects arising from the PA BCW(ms), and 2) forward DA flow is mainly sustained by reversal of PA flow in protodiastole but discharge of a more central reservoir in diastole.     

Testosterone Plus Low-Intensity Physical Training in Late Life Improves Functional Performance,Skeletal Muscle Mitochondrial Biogenesis,and Mitochondrial Quality Control in Male Mice

Testosterone supplementation increases muscle mass in older men but has not been shown to consistently improve physical function and activity. It has been hypothesized that physical exercise is required to induce the adaptations necessary for translation of testosterone-induced muscle mass gain into functional improvements. However, the effects of testosterone plus low intensity physical exercise training (T/PT) on functional performance and bioenergetics are unknown. In this pilot study, we tested the hypothesis that combined administration of T/PT would improve functional performance and bioenergetics in male mice late in life more than low-intensity physical training alone. 28-month old male mice were randomized to receive T/PT or vehicle plus physical training (V/PT) for 2 months. Compare to V/PT control, administration of T/PT was associated with improvements in muscle mass, grip strength, spontaneous physical movements, and respiratory activity. These changes were correlated with increased mitochondrial DNA copy number and expression of markers for mitochondrial biogenesis. Mice receiving T/PT also displayed increased expression of key elements for mitochondrial quality control, including markers for mitochondrial fission-and-fusion and mitophagy. Concurrently, mice receiving T/PT also displayed increased expression of markers for reduced tissue oxidative damage and improved muscle quality. Conclusion: Testosterone administered with low-intensity physical training improves grip strength, spontaneous movements, and respiratory activity. These functional improvements were associated with increased muscle mitochondrial biogenesis and improved mitochondrial quality control.     

Mitochondrial permeability transition mediates apoptosis induced by N-methyl(R)salsolinol,an endogenous neurotoxin,and is inhibited by Bcl-2 and rasagiline,N-propargyl-1(R)-aminoindan

The role of mitochondrial permeability transition (PT) in apoptosis induced by an endogenous neurotoxin, N-methyl(R)salsolinol [NM(R)Sal], was studied by use of dopaminergic neuroblastoma SH-SY5Y cells. NM(R)Sal reduced mitochondrial membrane potential, DeltaPsim, in the early phase of apoptosis, which was not suppressed by a pan-caspase inhibitor, but was antagonized by Bcl-2 and cyclosporin A, suggesting the involvement of the PT in NM(R)Sal-induced loss of DeltaPsim. NM(R)Sal-induced apoptosis was completely inhibited not only by Bcl-2 and a pan-caspase inhibitor, but also by cyclosporin A, suggesting the essential role of the PT in NM(R)Sal-induced apoptosis. In mitochondria isolated from rat liver, NM(R)Sal induced swelling and reduced DeltaPsim, which was inhibited by cyclosporin A and Bcl-2 overexpression. These results indicate that NM(R)Sal induced the PT by direct action on the mitochondria. Rasagiline, N-propargyl-1(R)-aminoindan, which is a now under a clinical trial for Parkinson's disease, suppressed the DeltaPsim reduction, release of cytochrome c, and apoptosis induced by NM(R)Sal in SH-SY5Y cells. Rasagiline also inhibited the NM(R)Sal-induced loss of DeltaPsim and swelling in the isolated mitochondria, proving that rasagiline directly targets the mitochondria also. Altogether, mitochondrial PT plays a key role both in NM(R)Sal-induced cell death and the neuroprotective effect of rasagiline.     

Two critical factors affecting the release of mitochondrial cytochrome C as revealed by studies using N,N'-dicyclohexylcarbodiimide as an atypical inducer of permeability transition

N,N'-dicyclohexylcarbodiimide (DCCD) was earlier reported to have stimulatory effects on mitochondrial respiration and to induce mitochondrial swelling, when it was added to mitochondrial suspensions. These data seem to imply that DCCD caused the mitochondrial permeability transition (PT), but this possibility had never been investigated. In the present study, effects of DCCD on the mitochondrial structure and function were studied in detail. DCCD was found to induce mitochondrial PT in a cyclosporine A-insensitive manner. Electron microscopic analysis also supported the induction of the mitochondrial PT by DCCD. However, different from many other PT inducers, DCCD failed to cause massive release of mitochondrial cytochrome c. To understand the relationship between the induction of mitochondrial PT and the release of mitochondrial cytochrome c, we compared the actions of DCCD on mitochondrial structure and function with those of Ca2+, known as an ordinary PT inducer. As a result, two parameters considered to be critical for controlling the release of mitochondrial cytochrome c on the induction of PT were mitochondrial volume and the velocity of mitochondrial oxygen consumption.     

Growth factors, signaling pathways, and the regulation of proliferation and differentiation in BC3H1 muscle cells. II. Two signaling pathways distinguished by pertussis toxin and a potential role for the ras oncogene

In the preceding report (Kelvin, D.J., G. Simard, H.H. Tai, T.P. Yamaguchi, and J.A. Connolly. 1989. J. Cell Biol. 108:159-167) we demonstrated that pertussis toxin (PT) blocked proliferation and induced differentiation in BC3H1 muscle cells. In the present study, we have used PT to examine specific growth factor signaling pathways that may regulate these processes. Inhibition of [3H]thymidine by PT in 20% FBS was reversed in a dose-dependent fashion by purified fibroblast growth factor (FGF). In 0.5% FBS, the normally induced increase in creatine kinase (CK) activity was blocked by FGF in both the presence and absence of PT. Similar results were obtained with purified epidermal growth factor (EGF). We subsequently examined the effect of a family of growth factors linked to inositol lipid hydrolysis and found that thrombin, like FGF, would increase [3H]thymidine incorporation and block CK synthesis. However, PT blocked thymidine incorporation induced by thrombin, and blocked the inhibition of CK turn-on in 0.5% FBS by thrombin. The ras oncogene, a G protein homologue, has previously been shown to block muscle cell differentiation in C2 muscle cells (Olson, E.N., G. Spizz, and M.A. Tainsky. 1987. Mol. Cell. Biol. 7:2104-2111); we have characterized a BC3H1 cell line, BCT31, which we transfected with the val12 oncogenic Harvey ras gene. This cell line did not express CK in response to serum deprivation. Whereas [3H]thymidine incorporation was inhibited by 70-80% by increasing doses of PT in control cells, BCT31 cells were only inhibited by 15-20%. ADP ribosylation studies indicate this PT-insensitivity is not because of the lack of a PT substrate in this cell line. Furthermore, PT could not induce CK expression in BCT31 cells as it did in parental cells. We conclude that there are at least two distinct growth factor pathways that play a key role in regulating proliferation and differentiation in BC3H1 muscle cells, one of which is PT sensitive, and postulate that a G protein is involved in transducing signals from the thrombin receptor. We believe that ras functions in the transduction of growth factor signals in the nonPT-sensitive pathway or downstream from the PT substrate in the second pathway.     

Apoptosis, cell proliferation and in vivo biological behaviour of primary and metastatic tumour cells of an AKR lymphoma variant

The possibility that apoptosis and/or cell proliferation have a role in tumour progression in a murine T cell lymphoma was tested. The model consisted of the comparison of primary (PT) and metastatic tumour (MT) cells. The PT cells, but not the MT cells displayed a very pronounced tendency for spontaneous apoptosis. Proliferative capacity of MT cells was lower than that of PT cells, suggesting that it does not contribute to the metastatic phenotype in this system. Release from apoptosis does however, probably, play a role in the aggressiveness of the lymphoma.