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PubMed1992

Motion of actin filaments in the presence of myosin heads and ATP.

We measured, by fluorescence correlation spectroscopy, the motion of actin filaments in solution during hydrolysis of ATP by acto-heavy meromyosin (acto-HMM). The method relies on the fact that the intensity of fluorescence fluctuates as fluorescently labeled actin filaments enter and leave a small sample volume. The rapidity of these number fluctuations is characterized by the autocorrelation function, which decays to 0 in time that is related to the average velocity of translation of filaments. The time of decay of the autocorrelation function of bare actin filaments in solution was 10.59 +/- 0.85 s. Strongly bound (rigor) heads slowed down the diffusion. Direct observation of filaments under an optical microscope showed that addition of HMM did not change the average length or flexibility of actin filaments, suggesting that the decrease in diffusion was not due to a HMM-induced change in the shape of filaments. Rather, slowing down of translational motion was caused by an increase in the volume of the diffusing complex. Surprisingly, the addition of ATP to acto-HMM accelerated the motion of actin filaments. The acceleration was the greatest at the low molar ratios of HMM:actin. Direct observation of filaments under an optical microscope showed that in the presence of ATP the average length of filaments did not change and that the filaments became stiffer, suggesting that acceleration of diffusion was not due to an ATP-induced increase in flexibility of filaments. These results show that some of the energy of splitting of ATP is impaired to actin filaments and suggest that 0.06 +/- 0.02 of HMM interferes with the diffusion of actin filaments during hydrolysis of ATP.

Actins↗
PubMed1993

Adiabatic compressibility of myosin subfragment-1 and heavy meromyosin with or without nucleotide.

The partial specific adiabatic compressibilities of myosin subfragment-1 (S1) and heavy meromyosin (HMM) of skeletal muscle in solution were determined by measuring the density and the sound velocity of the solution. The partial specific volumes of S1 and HMM were 0.713 and 0.711 cm3/g, respectively. The partial specific adiabatic compressibilities of S1 and HMM were 4.2 x 10(-12) and 2.9 x 10(-12) cm2/dyn, respectively. These values are in the same range as the most of globular proteins so far studied. The result indicates that the flexibility of S1 region almost equals to that of HMM. After binding to ADP.orthovanadate, S1 and HMM became softer than their complexes with ADP. The bulk moduli of S1 and HMM were of the order of (4-6) x 10(10) dyn/cm2, which are very comparable with the bulk modulus of muscle fiber.

Adenine Nucleotides↗
PubMed1988

Preparation and characterization of heavy meromyosin and subfragment 1 from vertebrate cytoplasmic myosins.

The soluble fragments of myosin, heavy meromyosin (HMM), and subfragment 1 (S-1) have been instrumental in elucidating the kinetic mechanisms of the actin-activated MgATPase activity of both skeletal and smooth muscle myosin. To date, relatively little has been published on these fragments from vertebrate cytoplasmic myosins. We now describe the preparation and steady-state kinetic characterization of S-1 and HMM from human platelet and avian intestinal epithelial brush border myosin. The HMM prepared from each of these tissues was similar both in their SDS-polyacrylamide gel pattern and in their steady-state kinetic properties. The Vmax of the actin-activated MgATPase activity varied between 0.8 and 2.5 s-1, and the KATPase (the apparent dissociation constant derived from a double-reciprocal plot of the MgATPase activity) was about 1-2 microM. This low value for the apparent dissociation constant was similar to the dissociation constant of HMM for actin directly measured under similar conditions and is about 40 times lower than that determined with avian smooth muscle HMM. The KATPase of the cytoplasmic HMM was only slightly increased when the ionic strength was raised from 12 to 112 mM.

Actins↗
PubMed2006

Phosphorylation of a single head of smooth muscle myosin activates the whole molecule.

Regulatory light chain (RLC) phosphorylation activates smooth and non-muscle myosin II, but it has not been established if phosphorylation of one head turns on the whole molecule. Baculovirus expression and affinity chromatography were used to isolate heavy meromyosin (HMM) containing one phosphorylated and one dephosphorylated RLC (1-P HMM). Motility and steady-state ATPase assays indicated that 1-P HMM is nearly as active as HMM with two phosphorylated heads (2-P HMM). Single-turnover experiments further showed that both the dephosphorylated and phosphorylated heads of 1-P HMM can be activated by actin. Singly phosphorylated full-length myosin was also an active species with two cycling heads. Our results suggest that phosphorylation of one RLC abolishes the asymmetric inhibited state formed by dephosphorylated myosin [Liu, J., et al. (2003) J. Mol. Biol. 329, 963-972], allowing activation of both the phosphorylated and dephosphorylated heads. These findings help explain how smooth muscles are able to generate high levels of stress with low phosphorylation levels.

Actins↗
PubMed1996

Polymerization and in vitro motility properties of yeast actin: a comparison with rabbit skeletal alpha-actin.

Actin purified from the yeast (Saccharomyces cerevisae) was polymerized faster than rabbit skeletal alpha-actin by MgCl2. The two actins polymerized at similar rates in the presence of CaCl2. Yeast actin, up to 25 microM, was not polymerized by KCl (100-300 mM); the monovalent salt also inhibited the MgCl2-induced polymerization of actin. The local structure of the subdomain-2 region in yeast actin filaments was probed by subtilisin and trypsin digestions. Loop 38-52 appeared more flexible and accessible to subtilisin in yeast than in rabbit actin. In contrast, tryptic digestions at Lys-61 and -68 occurred at the same rate for yeast and alpha-actin filaments. Modification of yeast actin by a sulfhydryl reagent CPM [7-(diethylamino)-3-(4'-maleimidophenyl)-4-methylcoumain] was specific to the Cys-374 residue; no labeling of a yeast actin mutant containing an alanine substitution for cysteine 374 was observed. The rates of Cys-374 labeling by CPM were similar for yeast and muscle actin, suggesting a similar environment for the C terminus in both polymers. In the in vitro motility assays, yeast actin required higher concentrations of heavy meromyosin (HMM) for its sliding than did the rabbit actin. At saturating concentrations of HMM, the sliding velocities of both actins were the same (3.0 microns/s). Relative forces generated by HMM with yeast and muscle actin were assessed by monitoring their in vitro motility in the presence of NEM-HMM load. The sliding of yeast actin was stopped at a level of external load (molar ratio NEM-HMM/HMM = 0.25) lower than that of muscle actin (NEM-HMM/HMM = 0.43), suggesting lower force production with yeast actin. These results are discussed in terms of the myosin cross-bridge cycle and actomyosin interactions.

Actins↗
PubMed1997

Activation of regulated actin by SH1-modified myosin subfragment 1.

The reactive SH1 (Cys-707) group of the myosin subfragment 1 (S1) has been used frequently as an attachment site for fluorescent and spin probes in solution and muscle fiber experiments. In this study we examined (i) the motor function of SH1 spin-labeled heavy meromyosin (HMM) in the in vitro motility assays and (ii) the effect of SH1-modified S1 on the motility of regulated actin, i.e., actin complexed with tropomyosin and troponin. N-ethylmaleimide (NEM), N-(1-oxyl-2,2,6,6-tetramethyl-4-piperidinyl)-iodacetamide (IASL), N-[[(iodoacetyl)amino]ethyl]1-sulfo-5-naphthylamine (IAEDANS), and iodoacetamide (IAA) were used to selectively modify the SH1 group on S1; the SH1 group on HMM was labeled with IASL. In the in vitro motility assays, 10-20% of unregulated actin filaments moved at a speed of approximately 1 microm/s over a surface coated with 90-95% modified IASL-HMM. Actin sliding was not observed with 95-98% modified IASL-HMM. The sliding of regulated actin over unmodified HMM was activated by the addition of S1 modified with any of the SH1 reagents to the in vitro motility assay solutions; both the speeds and the percentage of the moving filaments increased at pCa 5, 7, and 8. To shed light on the activation of regulated actin sliding by SH1-modifed S1, acto-S1 ATPase and the binding to actin were determined for IASL-S1. While the binding affinities to actin were similar for IASL-S1 and unmodified S1 in the presence and absence of ADP and ATP, the Km and Vmax values were approximately 10-fold lower for the modified protein. It is concluded that the activation of regulated actin by SH1-modifed S1 facilitates the interaction of unmodified HMM heads with actin and thus can increase the sliding speeds and the percentage of regulated actin filaments that move in the in vitro motility assays.

Actins↗
PubMed1997

Cooperativity and regulation of scallop myosin and myosin fragments.

Scallop heavy meromyosin (HMM) preparation obtained by a new improved method showed a Mg-ATPase activity that was activated 15-fold by calcium. The ATPase activity depended on ionic strength and reached maximum at 0.1 M without altering calcium sensitivity. The highly regulated HMM and myosin preparations showed cooperative properties not seen with unregulated subfragment 1 (S1). ATPase activity of myosin and HMM increased steeply with calcium concentration, yielding Hill coefficients about 3 and 4, respectively. Calcium binding by HMM and myosin became cooperative in the presence of ADP, AMP-PNP, or ADP.Vi yielding Hill coefficients of 1.8 and 2.8, respectively. Binding of calcium by HMM in the presence of ATP was also cooperative at physiological ionic strength, whereas at low ionic strength the data fit best to a simple binding curve. In contrast, calcium binding by unregulated S1 followed a normal binding curve and was not affected by the presence of nucleotide analogues. Calcium decreased the affinity of ADP and ADP-PNP to myosin and HMM, but had no effect on the nucleotide binding to S1. The results indicate that communication between the nucleotide and calcium binding sites requires the presence of two heads and exists only in the "off" state. We propose that in the presence of calcium, interaction between the two heads is disrupted and they act independently.

Adenosine Diphosphate↗
PubMed2001

Phalloidin affects the myosin-dependent sliding velocities of actin filaments in a bound-divalent cation dependent manner.

We examined sliding velocities in vitro of four types of actin filaments, that is, filaments with Ca2+ or Mg2+ bound at the high affinity metal binding site, each with rhodamine phalloidin bound with a high or low stoichiometry. When surfaces coated with a high density of heavy meromyosin (HMM) were used, high stoichiometric concentrations of rhodamine phalloidin reduced sliding velocities of only Ca2+-actin filaments, by 40%. As the HMM density on surfaces was reduced, continuous movement of actin filaments became dependent on the presence of methylcellulose and sliding velocities of all four types became progressively slower. Interestingly, Ca2+-actin filaments with a high stoichiometric concentration of rhodamine phalloidin were the fastest among the four types of filaments on sparse HMM surfaces. In contrast, phalloidin did not affect steady state ATPase activities of HMM in the presence of Ca2+- or Mg2+-actin filaments. We speculate that the reversal of the order of sliding velocities among the four types of actin filaments between high and low densities of HMM relates with different axial elasticity of the actin filaments, so that stiffer filaments move slower on dense HMM surfaces, but faster on sparse surfaces, than elastic ones.

Actin Cytoskeleton↗
PubMed1995

Functional chicken gizzard heavy meromyosin expression in and purification from baculovirus-infected insect cells.

The heavy chain and the essential and the regulatory light chains of chicken gizzard heavy meromyosin (HMM) were coexpressed in Spodoptera frugiperda (fall armyworm) cells infected with a mixture of two recombinant Autographa californica baculoviruses. Soluble HMM consisting of the heavy chain and the two types of light chains was obtained. The recombinant HMM was purified from the virus-infected cells and characterized. The regulatory light chain of the isolated recombinant HMM was phosphorylated by myosin light chain kinase in the presence of calmodulin in a Ca(2+)-dependent manner. The ATPase of the recombinant HMM was activated by rabbit skeletal muscle actin when myosin light chain kinase, calmodulin, and Ca2+ were present in the reaction medium. Chicken gizzard tropomyosin enhanced the actin-activated ATPase activity. The recombinant HMM decorated actin filaments, displaying the characteristic arrowhead pattern along the filaments. This report on a functional recombinant double-headed smooth muscle myosin fragment opens the way to detailed studies on the molecule.

Actins↗
PubMed1996

The role of surface loops (residues 204-216 and 627-646) in the motor function of the myosin head.

A characteristic feature of all myosins is the presence of two sequences which despite considerable variations in length and composition can be aligned with loops 1 (residues 204-216) and 2 (residues 627-646) in the chicken myosin-head heavy chain sequence. Recently, an intriguing hypothesis has been put forth suggesting that diverse performances of myosin motors are achieved through variations in the sequences of loops 1 and 2 [Spudich, J. (1994) Nature (London) 372, 515-518]. Here, we report on the study of the effects of tryptic digestion of these loops on the motor and enzymatic functions of myosin. Tryptic digestions of myosin, which produced heavy meromyosin (HMM) with different percentages of molecules cleaved at both loop 1 and loop 2, resulted in the consistent decrease in the sliding velocity of actin filaments over HMM in the in vitro motility assays, did not affect the Vmax, and increased the Km values for actin-activated ATPase of HMM. Selective cleavage of loop 2 on HMM decreased its affinity for actin but did not change the sliding velocity of actin in the in vitro motility assays. The cleavage of loop 1 and HMM decreased the mean sliding velocity of actin in such assays by almost 50% but did not alter its affinity for HMM. To test for a possible kinetic determinant of the change in motility, 1-N6-ethenoadenosine diphosphate (epsilon-ADP) release from cleaved and uncleaved myosin subfragment 1 (S1) was examined. Tryptic digestion of loop 1 slightly accelerated the release of epsilon-ADP from S1 but did not affect the rate of epsilon-ADP release from acto-S1 complex. Overall, the results of this work support the hypothesis that loop 1 can modulate the motor function of myosin and suggest that such modulation involves a mechanism other than regulation of ADP release from myosin.

Actins↗
PubMed1998

The shortening of the N-terminus of myosin essential light chain A1 influences the interaction of heavy meromyosin with actin.

The effects resulting from the removal of the N-terminus of heavy meromyosin (HMM) A1 light chain by papain digestion are investigated. The fluorometry of TRITC-phalloidin labelled actin in ghost fibers is used as a tool for sensing conformational changes of rigor complex of phosphorylated and dephosphorylated HMM with actin filament. The experiments were performed both under conditions assuring saturation of RLC with magnesium cation (4 mM EGTA) or calcium cation (0.1 mM CaCl2), and in constant presence of 1 mM magnesium chloride. HMM native and with A1 shortened from the N-terminus is used. As it was observed previously rigor complex of actin filament and native HMM shows sensitivity to the kind of cation saturating RLC and to the phosphorylation status of RLC. In particular, the sin2 theta parameter of actin bound rhodamine-phalloidin fluorescence polarization representing roughly the flexibility of actin filament HMM complex changes significantly with the changes of RLC phosphorylation and cation saturation. Removal of the N-terminus of A1 reduces this sensitivity to cation and phosphorylation both in the case of dephosphorylated and phosphorylated HMM. Our results suggest that the N-terminus of A1 plays significant role in the rigor interaction of myosin heads with actin and is involved in modulatory function of RLC in this interaction.

Actins↗
PubMed1990

Subtilisin cleavage of actin inhibits in vitro sliding movement of actin filaments over myosin.

Subtilisin cleaved actin was shown to retain several properties of intact actin including the binding of heavy meromyosin (HMM), the dissociation from HMM by ATP, and the activation of HMM ATPase activity. Similar Vmax but different Km values were obtained for acto-HMM ATPase with the cleaved and intact actins. The ATPase activity of HMM stimulated by copolymers of intact and cleaved actin showed a linear dependence on the fraction of intact actin in the copolymer. The most important difference between the intact and cleaved actin was observed in an in vitro motility assay for actin sliding movement over an HMM coated surface. Only 30% of the cleaved actin filaments appeared mobile in this assay and moreover, the velocity of the mobile filaments was approximately 30% that of intact actin filaments. These results suggest that the motility of actin filaments can be uncoupled from the activation of myosin ATPase activity and is dependent on the structural integrity of actin and perhaps, dynamic changes in the actin molecule.

Actin Cytoskeleton↗
PubMed1979

N-ethylmaleimide-modified heavy meromyosin. A probe for actomyosin interactions.

Treatment of rabbit skeletal muscle heavy meromyosin (HMM) with the sulfhydryl reagent N-ethylmaleimide (NEM) produces a species of HMM which remains tightly bound to actin in the presence of MgATP. NEM-HMM forms characteristic "arrowhead" complexes with actin which persist despite rinses with MgATP. NEM-HMM inhibits the actin activation of native HMM-ATPase activity, the superprecipitation of actomyosin, the contraction of glycerinated muscle myofibrils, and the contraction of cytoplasmic strands of the soil amoeba Chaos carolinensis. However, NEM-HMM does not interfere with in vitro microtubule polymerization or beating of demembranated cilia.

Actins↗
PubMed1981

Inhibition of palmitoyl CoA of EDTA- and Mg2+-ATPase of heavy meromyosin from rabbit skeletal muscle.

Palmitoyl CoA inhibited EDTA-ATPase of heavy meromyosin (HMM) prepared from rabbit skeletal muscle. The concentration for half maximum inhibition of EDTA-ATPase was about 18 microM. Myristoyl CoA, the other long chain fatty acyl CoA, also inhibited EDTA-HMM ATPase, but CoA and short chain CoA thioesters, such as butyryl CoA, acetoacetyl CoA and acetyl CoA, at 40 microM hardly inhibited EDTA-ATPase. Less than 20% inhibition of EDTA-HMM ATPase was obtained with Na-palmitate and Na-myristate at 40 microM, whereas about 90% inhibition of the enzyme occurred in the presence of 40 microM palmitoyl CoA and myristoyl CoA. Palmitoyl carnitine, as well as carnitine, failed to inhibit EDTA-HMM-ATPase. The inhibition of palmitoyl CoA of EDTA-ATPase was reversed by bovine serum albumin and spermine. Mg2+-HMM ATPase activity was enhanced by palmitoyl CoA at 2, 5, and 10 microM. About a 25% increase in Mg2+-HMM ATPase activity was obtained at 5 and 10 microM. At higher concentrations than 20 microM, the enzyme was inhibited by palmitoyl CoA and the degree of inhibition was related to the concentration of the CoA thioester. At 80 microM, the activity was about 15% of the maximum value. The efficacy of myristoyl CoA on Mg2+-ATPase was almost the same as that of palmitoyl CoA. Mg2+-ATPase activity was not enhanced by CoA, butyryl CoA, acetoacetyl CoA, Na-myristate, Na-palmitate, palmitoyl carnitine, or carnitine at 10 microM, and was hardly reduced by these substances at 40 microM. Serum albumin and spermine also canceled, to some extent, these effects of palmitoyl CoA on Mg2+-ATPase.

Acyl Coenzyme A↗
PubMed1985

Relationship between the ATPase activity and the ATP-induced fluorescence enhancement of SH-modified heavy meromyosin during its fractional inactivation by vanadate plus ADP: evidence for heterogeneity in the active sites.

We have examined whether heavy meromyosin (HMM) consists of a single kind of active site by analyzing the changes in the relative MgATPase activity and the relative amplitude of the ATP-induced fluorescence enhancement of the protein when the fraction of HMM "affinity"-labeled by vanadate plus ADP was varied. The analysis is based on a prediction that these two changes should be proportional to each other if myosin consists of a single kind of active site and generates the rate-limiting myosin**product complex emitting enhanced fluorescence. Although the difference between these two changes was very small with native HMM, it was large with HMM in which 5 fast-reactive sulfhydryl-groups per head were pre-modified with thimerosal. The difference indicated the existence of heterogeneous active sites in the SH-modified HMM. The results were best explained in terms of the hypothesis that fifty percent of the active site splits MgATP by a mechanism giving a fluorescence enhancement whereas the other fifty percent splits MgATP by another mechanism giving no fluorescence enhancement. Two possible explanations for the existence of heterogeneous active sites in the SH-modified HMM are discussed. One assumes the pre-existence of some sort of 1:1 heterogeneity in the micro-environment of the active sites and the other, which is considered less likely, assumes the introduction of the heterogeneity as a result of the SH-modification.

Actins↗
PubMed1995

In vivo antitumor activity of hexamethylmelamine against human breast, stomach and colon carcinoma xenografts.

We have evaluated the antitumor activity of Altretamine (hexamethylmelamine, HMM) on human carcinoma xenografts serially transplanted in nude mice. Five human breast carcinoma xenografts, MX-1, T-61, MCF-7, R-27 and Br-10, were inoculated subcutaneously into female nude mice. Two human stomach carcinoma xenografts, SC-1-NU and St-4, and three human colon carcinoma xenografts, Co-3, Co-4 and Co-6, were inoculated subcutaneously into male nude mice. One pellet of 17 beta-estradiol (0.1 mg/mouse) was inoculated subcutaneously in the mice transplanted with MCF-7 when the tumors were inoculated. HMM was administered per os daily for 4 weeks. MX-1 and T-61 tumors regressed completely after treatment with HMM at a dose of 75 mg/kg (the maximum tolerated dose: MTD) for MX-1 and 25 mg/kg for T-61. Br-10 was sensitive, whereas MCF-7 and R-27 were resistant to HMM at its MTD. HMM exerted the most potent antitumor effect against T-61. Against MX-1, it exerted an antitumor effect equivalent to that of cisplatin or cyclophosphamide. In addition, this agent was effective against all stomach and colon carcinoma xenografts, in particular St-4 (T/C% = 10.7: the mean tumor weight of treated group/the mean tumor weight of control group) and Co-3 (T/C% = 31.5%) which are insensitive to presently available agents. HMM seems worthy of further clinical investigation as a candidate agent to treat breast, stomach, colon and other carcinomas.

Altretamine↗
PubMed2006

Intermonomer cross-linking of F-actin alters the dynamics of its interaction with H-meromyosin in the weak-binding state.

Previous cross-linking studies [Kim E, Bobkova E, Hegyi G, Muhlrad A & Reisler E (2002) Biochemistry 41, 86-93] have shown that site-specific cross-linking among F-actin monomers inhibits the motion and force generation of actomyosin. However, it does not change the steady-state ATPase parameters of actomyosin. These apparently contradictory findings have been attributed to the uncoupling of force generation from other processes of actomyosin interaction as a consequence of reduced flexibility at the interface between actin subdomains-1 and -2. In this study, we use EPR spectroscopy to investigate the effects of cross-linking constituent monomers upon the molecular dynamics of the F-actin complex. We show that cross-linking reduces the rotational mobility of an attached probe. It is consistent with the filaments becoming more rigid. Addition of heavy meromyosin (HMM) to the cross-linked filaments further restricts the rotational mobility of the probe. The effect of HMM on the actin filaments is highly cooperative: even a 1 : 10 molar ratio of HMM to actin strongly restricts the dynamics of the filaments. More interesting results are obtained when nucleotides are also added. In the presence of HMM and ADP, similar strongly reduced mobility of the probe was found than in a rigor state. In the presence of adenosine 5'[betagamma-imido] triphosphate (AMPPNP), a nonhydrolyzable analogue of ATP, weak binding of HMM to either cross-linked or native F-actin increases probe mobility. By contrast, weak binding by the HMM/ADP/AlF4 complex has different effects upon the two systems. This protein-nucleotide complex increases probe mobility in native actin filaments, as does HMM + AMPPNP. However, its addition to cross-linked filaments leaves probe mobility as constrained as in the rigor state. These findings suggest that the dynamic change upon weak binding by HMM/ADP/AlF4 which is inhibited by cross-linking is essential to the proper mechanical behaviour of the filaments during movement.

Actins↗
PubMed1998

The effect of recombinant TGFalpha, human milk, and human milk macrophage media on gut epithelial proliferation is decreased in the presence of a neutralizing TGFalpha antibody.

OBJECTIVE: An in vitro model was devised to compare the relative effects of recombinant transforming growth factor-alpha (TGFalpha), aqueous human milk, and human milk macrophage (HMM) medium on human fetal small intestinal cell (FHs-74) proliferation. METHODS: Recombinant TGFalpha at increasing concentrations (range 0.01-1,000 ng/ml media), the aqueous fraction of human milk (AHM), or HMM medium was added to FHs-74 cells in the presence or absence of a neutralizing TGFalpha antibody (1 microgram/ml medium). At 24 h, cell proliferation was measured and expressed as percent control. The experimental variables were (1) activators of cell growth (TGFalpha, AHM, and HMM medium); (2) increasing concentrations of TGFalpha, and (3) neutralizing antibody to TGFalpha. The dependent variable for all experiments was cell proliferation. RESULTS: Significant effects for growth stimulators and TGFalpha concentration as measured by cell proliferation were found. Specifically, there was a dose-dependent effect of TGFalpha on cell proliferation to the 5-ng/ml concentration, with a plateau reached in cell proliferation at higher concentrations. The stimulatory effect of TGFalpha was decreased in the presence of TGFalpha antibody (mean +/- SD 22 +/- 7. 1% decline, p < 0.001). In the presence of TGFalpha antibody, there was a 25 +/- 3.1% decline in HM-stimulated growth (p < 0.004), and a 27.6 +/- 3.2% decline in HMM medium-stimulated growth (p < 0.001). CONCLUSIONS: Neutralization of recombinant TGFalpha and that present in human milk and HMM medium by TGFalpha antibody led to a consistent decrease in in vitro human fetal small intestine epithelial proliferation without affecting cell viability. These results support the hypothesis that TGFalpha, whether derived from human recombinant sources, human milk or HMM medium has a measurable, trophic effect on in vitro human gut epithelial cells.

Animals↗