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Reduction in experimental pericardial adhesions using a hyaluronic acid bioabsorbable membrane.

Various materials have been suggested for pericardial closure following cardiac surgery to limit postoperative adhesion formation. Unfortunately, these devices have caused difficulties with infection, calcification, and epicardial scar formation. A bioabsorbable membrane with a short in-vivo persistence time, acting as a barrier to adhesion formation until mesothelial healing occurs, might obviate many of these problems. We examined the efficacy of a bioabsorbable membrane containing hyaluronic acid in the prevention of experimental pericardial adhesions; hyaluronic acid solutions have been shown experimentally to reduce adhesions following abdominal, orthopedic and cardiac surgery. Eleven dogs underwent sternotomy and pericardiotomy followed by 2-h protocol of desiccation and abrasion of pericardial and epicardial surfaces. In five dogs, the epicardial surfaces were covered with the test material, followed by pericardial and sternotomy closure. Six dogs served as controls. All animals underwent resternotomy 8 weeks after surgery; intrapericardial adhesions were graded on a severity scale (0-4). The test animals (mean adhesion score 1.4 +/- 0.5) had either no adhesions or filmy, avascular adhesions easily taken down with blunt dissection. In contrast, dense adhesions were found in the control animals (mean score 3.2 +/- 0.4), frequently obscuring tissue planes and epicardial anatomy. Use of the bioabsorbable membrane significantly reduced adhesion formation (test vs control, P < 0.001, Mann-Whitney test). We conclude that a hyaluronic acid membrane reduces postoperative pericardial adhesions in this canine model, and that the use of a bioabsorbable barrier in the prevention of pericardial adhesions warrants further investigation.

Animals↗

Hyaluronan mediates adhesion of metastatic colon carcinoma cells.

BACKGROUND: Hyaluronan (HA) is a cell-surface glycosaminoglycan that has been implicated in cancer progression. Cells isolated from metastatic colon carcinoma (SW620) produce greater amounts of pericellular HA than cells isolated from a primary tumor (SW480). Inhibition of hyaluronan synthases (HAS) by transfection with antisense cDNA decreases HA production. Because adhesion to the extracellular matrix (ECM) is required for invasion and metastasis, we hypothesized that pericellular HA mediates adhesion to ECM proteins such as laminin, collagen, and fibronectin and that inhibition of HA production or removal of HA by digestion with hyaluronidase would impair adhesion. MATERIALS AND METHODS: SW480, SW620, and antisense transfectants (SW620 cells transfected with vector alone, antisense HAS2, antisense HAS3, and both antisense HAS2 and HAS3) were assessed for adhesion to laminin, Type 1 collagen, or fibronectin-coated plates. To confirm that adhesion was mediated by HA, cells were treated with or without hyaluronidase prior to the assays. RESULTS: Metastatic SW620 cells adhered well to laminin; SW480 cells demonstrated 46% less adhesion (P < 0.05; Student's t test). SW620 cell adhesion to Type 1 collagen and fibronectin was >50% less than adhesion to laminin. Inhibition of HAS2 and/or HAS3 or pretreatment with hyaluronidase significantly decreased adhesion of SW620 cells to laminin (P < 0.05), suggesting that adhesion was dependent upon pericellular HA. CONCLUSIONS: Metastatic SW620 cells that produce large amounts of pericellular HA adhered well to laminin. Inhibition of HAS2 and/or HAS3 expression, or hyaluronidase digestion of pericellular HA significantly inhibited adhesion. These data suggest that HA promotes adhesion to laminin and may thereby facilitate invasion of the basement membrane and metastasis in colon carcinoma.

Carcinoma↗

Corneal epithelial adhesion abnormalities associated with LASIK.

PURPOSE: To assess the clinical characteristics, incidence, and pathologic correlation of corneal epithelial adhesion abnormalities encountered during LASIK. DESIGN: Prospective noncomparative interventional case series. PARTICIPANTS: Five hundred consecutive eyes of 268 patients undergoing primary LASIK procedures by one surgeon utilizing the Moria LSK One microkeratome and VISX Star S-2 excimer laser. METHODS: Corneal epithelial adhesion was assessed immediately preoperatively using a cellulose surgical sponge (adhesion test), and the incidence, extent, and location of epithelial defects occurring during the microkeratome incision of the corneal flap were recorded. Epithelial specimens from 7 corneas requiring debridement of dysadhesive epithelium were examined by transmission electron microscopy. MAIN OUTCOME MEASURES: The characteristics of the study population (age, gender, contact lens use, relevant ocular surface or systemic disease, refractive error, keratometry, pachymetry) and the microkeratome-related variables (head selection and vacuum level) were compared with the results of the preoperative epithelial adhesion test, the development of intraoperative epithelial effects (size and location), and the postoperative outcome. RESULTS: Epithelial defects were sustained by 51 corneas (10.2%), and among these, 31 (6.2%) were microdefects and 20 (4.0%) were macrodefects. The adhesion test was positive (indicative of compromised adhesion of epithelium to stroma) in 20 (64.5%) corneas with microdefects, but false negatives (epithelial defect despite negative adhesion test) occurred in 11 cases (35.4%). The adhesion test was positive in 16 (80%) of corneas having macrodefects, with 4 (20%) false negatives. Thus, the overall positive predictive value of the adhesion test was 59%, and the percentage of positive prediction was 92% (Bayes' theorem). Among all other outcome measures assessed, only corneal flap thickness seemed a potential risk factor, as 40 (78.4%) epithelial defects were associated with the creation of a 180- micro m-thick flap, whereas 10 (19.6%) were associated with a 160- micro m-thick flap and only 1 (2%) occurred with a 130- micro m flap. These trends were not, however, statistically significant (P = 0.15, Fisher exact test). Transmission electron microscopy of all epithelial debridement specimens disclosed consistent abnormality of the basement membrane adhesion complex, as thickened and multilaminated basement membrane remained adherent to the intact epithelial sheet. CONCLUSIONS: : Corneal epithelial dysadhesion and defects occurring in the course of LASIK surgery may be associated with an intrinsic compromise of the basement membrane adhesion complex, as evidenced clinically by the adhesion test and demonstrated pathologically by duplicated basement membrane.

Adult↗

Measurement of adhesion of human platelets in plasma to protein surfaces in microplates.

INTRODUCTION: Platelet adhesion is an initial, crucial and complex event for inhibiting blood loss upon vascular injury. Activation and adhesion of platelets also play a fundamental role in the development of thrombosis. A combination of exposed extracellular matrix proteins in the vascular wall and release of activating compounds from the participating cells activate the platelets. New potent anti-platelet agents are in progress but there is a shortage of methods that measure the concerted action of adhesive surfaces and soluble compounds upon platelet adhesion in vitro. The aim of this work was to develop a method to measure adhesion of platelets in plasma with standard laboratory equipment. METHODS: Platelet-rich plasma from healthy humans was used in studies to optimise the conditions of the present assay. Different proteins were coated in microplate wells and various soluble platelet activators and inhibitors were added to establish the ability of the current method to detect increased as well as decreased platelet adhesion. The amount of platelet adhesion was measured by the reaction between p-nitrophenyl phosphate and the intracellular enzyme acid phosphatase. RESULTS: Adhesion of platelets in plasma to microplate wells coated with albumin, collagen, fibrinogen and activated plasma showed significant surface dependency. The known soluble platelet activators adenosine diphosphate, adrenaline and ristocetin enhanced the levels of adhesion. Available anti-platelet agents such as prostacyclin, forskolin, acetylsalicylic acid and RGD containing peptides caused dose-dependent inhibition of platelet adhesion. DISCUSSION: This report describes a further development of a previously described method and offers the advantage to use platelets in plasma to measure platelet adhesion to protein surfaces. The assay is simple and flexible and is suitable in basic research for screening and characterisation of platelet adhesion responsiveness.

Adenosine Diphosphate↗

Usage of denture adhesives.

OBJECTIVES: There have been few reports in the dental literature on the number of denture wearers who regularly use denture adhesive. The objectives of this study were to see what the incidence of usage of denture adhesive was by a surveyed edentulous population, to determine the degree of success of its usage and the reasons for its current use or nonuse. METHODS: In this study, 146 patients attending the Adelaide Dental Hospital for denture treatment were surveyed regarding their usage of denture adhesive, using a prepared questionnaire. The surveyed group was divided into three categories--those who had never tried denture adhesive, those who had tried denture adhesive but no longer used it, and those who currently used denture adhesive. RESULTS: In the survey, there were 52 males (35.6%) and 94 females (64.4%) and 96 (65.7%) were over 60 years of age. Of these 78 patients (54.9%) had worn their dentures for 10 years or more, and 26 (17.8%) for 20 years or more. A total of 98 patients (67.1%) had never tried denture adhesive: 48 patients (32.9%) had tried denture adhesive but only 10 (6.9%) currently used it. Various reasons for trying denture adhesive and for its continued use or nonuse were given. CONCLUSIONS: In this study, 52% of patients surveyed saw no need for the use of denture adhesive as they managed their dentures well; 20.5% of patients did not know that denture adhesives existed; and 32.9% had tried denture adhesive but only 6.9% continued to use it on a regular basis.

Adhesives↗

[Interest of auto-cross-linked hyaluronic acid gel in the prevention of intrauterine adhesions after hysteroscopic surgery: A case-control study].

OBJECTIVES: To evaluate the efficacy of auto-cross-linked hyaluronic acid gel in the prevention of adhesions after operative hysteroscopy using a case-control study. MATERIALS AND METHODS: Fifty-four patients with an intrauterine lesion (myoma, polyp, uterine septa and adhesions) undergoing hysteroscopic surgery were divided into two groups: group A (30 patients) with intrauterine application of hyaluronic acid gel at the end of the surgical procedure and group B, which was considered as control (24 patients). The rate of adhesion formation, the score and the adhesion severity were estimated for each group using American Fertility Society (AFS) classification, by diagnostic hysteroscopy two months after surgery. No other treatment was associated. RESULTS: Age, weight, parity, hysterometry were comparable in the two groups. Surgery indications were polyp(s), myoma(s), uterine septa, and adhesions (11, 8, 1, and 10 patients in group A and 6, 6, 4, and 8 patients in group B, respectively). No difference was observed in intrauterine adhesion formation between the two groups (33.3% for group A and B; p = NS). The median adhesion scores using AFS were comparable in the two groups (1.30+/-2.35 vs 1.42+/-2.47; respectively, p = NS). The severity of the adhesions showed no significant difference between the two (70% stage I, mild adhesions; 20% stage II, moderate adhesions; 10% stage III, severe adhesions and 62.5% stage I; 25% stage II; 12.5% stage III in the group A and B, respectively). No adverse effect with the ACP gel was detected. CONCLUSION: ACP gel does not reduce the incidence and the severity of intrauterine adhesions after hysteroscopic surgery.

Adult↗

Insulin like growth factor-1 activates nuclear factor-kappaB and increases transcription of the intercellular adhesion molecule-1 gene in endothelial cells.

A critical early event in the pathogenesis of occlusive vascular disease is the adhesion of monocytes to endothelial cells. The authors have previously reported that insulin-like growth factor-1 increases monocyte-endothelial cell adhesion and increases the expression of intercellular adhesion molecule-1. In this study, it is hypothesized that the upregulation of intercellular adhesion molecule-1 expression after treatment with insulin-like growth factor-1 is caused by an increase in the transcription of intercellular adhesion molecule-1 in endothelial cells, and that this transcription is regulated, at least in part, by activation of nuclear factor-kappaB. Adherence cell assays were performed using insulin-like growth factor-1 treated human umbilical vein endothelial cells and human monocytes. To determine the role of nuclear factor-kappaB, Western blotting using the anti-p65 (activated portion of nuclear factor-kappaB) was performed on cell lysate of human umbilical vein endothelial cells treated with insulin-like growth factor-1. RT-PCR was performed on RNA extracted from insulin-like growth factor-1-treated human umbilical vein endothelial cells. Intercellular adhesion molecule-1 antibody attenuated the increase in monocyte-endothelial cell adhesion of endothelial cells exposed to insulin-like growth factor-1. We observed an increase in expression of the activated nuclear factor-kappaB p65 protein in response to insulin-like growth factor-1 treatment. Peak increase occurred at 30 min. This effect was sensitive to pretreatment of human umbilical vein endothelial cells with the insulin-like growth factor-1 receptor antibody. Human umbilical vein endothelial cells treated with insulin-like growth factor-1 for 2 and 4 h revealed a significant increase in intercellular adhesion molecule-1 mRNA as compared with untreated human umbilical vein endothelial cells. Tumor necrosis factor-alpha produced a larger increase in intercellular adhesion molecule-1 mRNA expression. These results suggest that insulin-like growth factor-1 enhances intercellular adhesion molecule-1 transcription and activates nuclear factor-kappaB in endothelial cells. The intracellular pathways that increase cell adhesion molecule expression may provide a common link to understanding the monocyte-endothelial cell adhesion that occurs in the early stages of atherosclerosis and restenosis.

Arteriosclerosis↗

Adhesion properties of enterococci to intestinal mucus of different hosts.

The adhesive capacity of selected enterococci to human, canine and porcine intestinal mucus was investigated in order to select for potential probiotic strains with good adhesive properties for human or animal use. The adhesion to the human intestinal mucus of the tested strains was found to range from log10 3.8 to log10 8.6 cfu per microtitre plate well. The highest adhesion to the human intestinal mucus was found among strains from horse faeces, dog faeces and dog feed. The adhesion to canine mucus was observed to range from log10 3.8 to log10 8.3 cfu/well, with the highest adhesive capacity among strains from dog faeces, horse faeces and dog's feed; on average log10 7.9, 7.3 and 7.0 cfu/well, respectively. Isolates from dogs did not bind at higher levels to canine mucus than to human mucus. A strong correlation was observed for the adhesion to human and canine intestinal mucus (p < 0.0001) and also between porcine and canine or human mucus (p < 0.05 for both). When comparing the adhesion of Enterococcus faecium and E. faecalis, no statistical significant differences were observed for any of the tested mucus types. The tested Enterococcus strains were found to exhibit a strain dependent on in vitro adhesion to human, canine and porcine intestinal mucus and did not exhibit host specificity in their adhesion, though some sources appeared to contain more adhesive strains than others. To our knowledge, this is the first report on the in vivo adhesion to intestinal mucus of a large number of enterococci from different sources.

Animal Feed↗

The effects of heparin and related molecules upon the adhesion of human polymorphonuclear leucocytes to vascular endothelium in vitro.

1. The effects of an unfractionated heparin preparation (Multiparin), a low molecular weight heparin preparation (Fragmin) and a selectively O-desulphated derivative of heparin lacking anticoagulant activity, have been investigated for their effects on the adhesion of human polymorphonuclear leucocytes (PMNs) to cultured human umbilical vein endothelial cells (HUVECs) in vitro. The effect of poly-L-glutamic acid, a large, polyanionic molecule was also studied. 2. Unfractionated heparin (50-1000 U ml-1), the O-desulphated derivative (0.3-6 mg ml-1) and the low molecular weight heparin (50 U-1000 U ml-1) all inhibited significantly the adhesion of 51Cr labelled PMNs to HUVECs stimulated with interleukin-1 beta (IL-1 beta; 10 U ml-1), bacterial lipopolysaccharide (LPS; 2.5 micrograms ml-1) or tumour necrosis factor-alpha (TNF-alpha; 125 U ml-1) for 6 h, whereas poly-L-glutamic acid had no effect. In addition, the three heparin preparations in the same concentration range inhibited significantly the adhesion of f-met-leu-phe-stimulated PMNs to resting HUVECs. 3. The effects of unfractionated heparin upon the expression of adhesion molecules intercellular adhesion molecule-1 (ICAM-1) and E-selection were also investigated, as were the effects of unfractionated heparin upon adhesion of human PMNs to previously stimulated HUVECs. Heparin had little effect upon levels of expression of these adhesion molecules on stimulated HUVECs. However, a profound effect upon PMN adhesion to previously stimulated HUVECs was demonstrated using the same preparation, suggesting that inhibition of adhesion molecule expression is not a major component of the described inhibitory effects of heparin. 4. Pre-incubation of PMNs with heparin followed by washing inhibited their adhesion to HUVECs, under different conditions of cellular activation, implying that heparin can bind to these cells and exert its anti-adhesive effects even when not directly present in the system. 5. These observations would suggest that both heparin and a low molecular weight heparin are capable of inhibiting adhesion of human PMNs to endothelial cells, an effect not dependent solely upon the polyanionic nature of these molecules, nor dependent upon their ability to act as anticoagulants.

Anticoagulants↗

Tissue markers as predictors of postoperative adhesions.

BACKGROUND: Postoperative adhesion formation has been associated with a decreased capacity to degrade intra-abdominally deposited fibrin. Adhesions, once lysed, have a high propensity for reformation. This study tested the hypothesis that patients with a high propensity for adhesion formation as well as adhesion tissue had a reduced fibrinolytic capacity. METHODS: Peritoneal biopsies were taken during abdominal surgery from 21 patients who had previously undergone operation; previously formed adhesion tissue was sampled from ten of these patients. Adhesion formation was scored. The fibrinolytic capacity of peritoneum was determined in tissue extracts. RESULTS: At the time of opening of the abdominal cavity, levels of plasminogen activator inhibitor (PAI) type 1 (P = 0.009) and tissue-type plasminogen activator (tPA)/PAI complex (P = 0.008) were increased in peritoneal samples from patients with severe adhesions compared with those in samples from patients with less severe adhesions. Adhesion tissue similarly had reduced fibrinolytic capacity as judged by a decrease in tPA activity (P = 0.005) and an increase in PAI-1 level (P = 0.01), reflected in an increased level of tPA/PAI complex (P = 0.008) compared with unaffected peritoneum. CONCLUSION: These observations demonstrate reduced fibrinolytic capacity in peritoneal tissue in patients with a greater propensity for development of adhesions and likewise in adhesion tissue. This suggests that components of the fibrinolytic system may be used as markers of an increased risk of adhesion development.

Biomarkers↗

Wound glue: a comparative study of tissue adhesives.

UNLABELLED: The purpose of this study was to determine which single-use wound adhesive is the most appropriate in terms of ease of use, minimal pain on application, adequate bonding time and wound closure. The three wound adhesives audited were Indermil (n-butyl cynoacrylate), Liquiband (n-butyl cynoacrylate) and Dermabond (octylcyanoacrylate). SAMPLE AND SETTING: The study was conducted in an urban paediatric emergency department treating over 39,000 patients annually. The sample was taken from the client population presenting with lacerations requiring tissue adhesive closure, within the limitations of the study (n = 63). METHODOLOGY: A non-blinded comparative study was performed. Children presenting with an appropriate laceration were assigned to receive either Indermil, Dermabond or Liquiband. The wounds were closed following the guidelines stated by the individual manufacturers. The nurses administering the tissue adhesive were asked to complete the audit form post closure and to comment on the procedure in descriptive terms. RESULTS: Scalp wounds accounted for 79% (n = 50) of all the lacerations closed in the study. None of the glues were reported to be completely pain-free. However, the Liquiband tissue adhesive produced an average pain score of only 0.1, whereas the Dermabond tissue adhesive scored the highest at 0.97. The nurses using the tissue adhesives reported that Liquiband was the best tissue adhesive in terms of wound closure and ease of use. However, the only tissue adhesive to report a 100% success rate was Indermil. DISCUSSION AND RECOMMENDATIONS: All of the tissue adhesives examined produced satisfactory results in terms of wound closure and ease of use. However, the Liquiband tissue adhesive produced the most consistent results, scoring higher in most of the categories when compared with the other tissue adhesives.

Attitude of Health Personnel↗

Minimal adhesions to ePTFE mesh after laparoscopic ventral incisional hernia repair: reoperative findings in 65 cases.

BACKGROUND AND OBJECTIVES: Laparoscopic ventral incisional hernia repair involves intraabdominal placement of a synthetic mesh, and the possibility of formation of severe visceral adhesions to the prosthesis is a principal concern. Little clinical information based on reoperative findings is available about adhesions to biomaterials placed intraabdominally. We conducted a multi-institutional study of adhesions to implanted expanded polytetrafluoroethylene (ePTFE) mesh at reoperation in patients who had previously undergone laparoscopic incisional hernia repair done with the same mesh implantation technique. METHODS: Nine surgeons retrospectively assessed the severity of adhesions to ePTFE mesh at reoperation in 65 patients. For each case, adhesions were assigned a score of 0 to 3, with 0 indicating no adhesions and 3 severe adhesions. RESULTS: The mean time from mesh implantation to reoperation was 420 days (range, 2-1 739 days). No adhesions were observed in 15 cases. Forty-four cases received an adhesion score of 1, and 6 cases a score of 2; no scores of 3 were assigned. Thus, 59 patients (91 %) had either no or filmy, avascular adhesions. No enterotomies occurred during adhesiolysis. CONCLUSIONS: In this large series of reoperations after laparoscopic incisional hernia repair, no or minimal formation of adhesions to implanted ePTFE mesh was observed in 91 % of cases, and no severe cohesive adhesions were found. Comparative analyses of newer materials based on clinical reoperative findings are warranted to assess the safety of intraabdominally placed meshes.

Adult↗

[O44--a solvent for silicone oil adhesions on intraocular lenses].

AIM: To examine the efficiency of O44, a partial fluorinated octane, as a solvent for silicone oil adhesions on intraocular lenses. MATERIALS AND METHODS: After placing silicone- and PMMA-lenses in silicone oil, the adhesions were treated with O44. The extent of silicone oil adhesions and the effectiveness of O44 were studied by gross microscopy by scanning electrone microscopy (SEM) and combined energy dispersive spectrometry (EDX). Furthermore an explanted silicone lens with oil adhesions was treated with O44 and examined. To simulate the effect of adhesion proteins PMMA- and silicone lenses were placed in silicone oil and human plasma. RESULTS: With EDX it was possible to prove that O44 is able to remove silicone oil adhesions from PMMA-lenses. The removal from an explanted silicone lens by O44 could be demonstrated by light microscopy. Silicone oil adhesions on intraocular lenses (IOL's) in vitro showed a different oil coverage than the IOL in vivo. Silicone lenses in vitro were often covered by a homogenous oil film and therefore the demonstration of the silicone adhesions and of the efficacy of O44 was difficult. The adhesions of IOL's placed additional in human plasma did not show any differences. CONCLUSIONS: O44 may be a successful intraoperative tool to remove silicone oil adhesions avoiding explantation of silicone oil contaminated IOL's. Silicone oil adhesions in vivo seem to be influenced by adhesive proteins.

Efficiency↗

The minimal essential unit for cadherin-mediated intercellular adhesion comprises extracellular domains 1 and 2.

N-cadherin comprises five homologous extracellular domains, a transmembrane, and a cytoplasmic domain. The extracellular domains of N-cadherin play important roles in homophilic cell adhesion, but the contribution of each domain to this phenomenon has not been fully evaluated. In particular, the following questions remain unanswered: what is the minimal domain combination that can generate cell adhesion, how is domain organization related to adhesive strength, and does the cytoplasmic domain serve to facilitate extracellular domain interaction? To address these issues, we made serial constructs of the extracellular domains of N-cadherin and produced various cell lines to examine adhesion properties. We show that the first domain of N-cadherin alone on the cell surface fails to generate adhesive activity and that the first two domains of N-cadherin form the "minimal essential unit" to mediate cell adhesion. Cell lines expressing longer extracellular domains or N-cadherin wild type cells formed larger cellular aggregates than those expressing shorter aggregates. However, adhesion strength, as measured by a shearing test, did not reveal any differences among these aggregative cell lines, suggesting that the first two domains of N-cadherin cells generate the same strength of adhesive activity as longer extracellular domain cells. Furthermore, truncations of the first two domains of N-cadherin are also sufficient to form cisdimerization at an adhesive junction. Our findings suggest that the extracellular domains of N-cadherin have distinct roles in cell adhesion, i.e. the first two domains are responsible for homophilic adhesion activity, and the other domains promote adhesion efficiency most likely by positioning essential domains relatively far out from the cell surface.

Animals↗

In vitro study of transdermal nicotine delivery: influence of rate-controlling membranes and adhesives.

The objective of this study was to evaluate the influence of a rate-controlling membrane and adhesive on the in vitro permeation of nicotine. The physicochemical properties of the adhesive, including adhesion and rheology (viscosity), were also detected. Higher permeability of nicotine was observed through a hydrophilic membrane than through a hydrophobic membrane. Natural rubber and silicone were used as the adhesive bases, respectively. The silicone adhesive showed the highest adhesion among all adhesive formulations. To increase the adhesion of natural rubber, a tackifier (polyisoprene) and a secondary tackifier (terpene polymer; Px 1150) were incorporated into the formulations to achieve acceptable adhesion. The nicotine permeation through silicone adhesive and three natural rubber adhesives with the secondary tackifier (2%, 4%, and 6% Px 1150) was close to that from a commercially available patch (Habitrol), although the loading amount of nicotine was not the same. A longer lag time during the in vitro permeation study of nicotine was required for the adhesives prepared in our laboratory than for the commercially available patch.

Adhesives↗

The role of integrins alpha 2 beta 1 and alpha 3 beta 1 in cell-cell and cell-substrate adhesion of human epidermal cells.

We have examined cultures of neonatal human foreskin keratinocytes (HFKs) to determine the ligands and functions of integrins alpha 2 beta 1, and alpha 3 beta 1 in normal epidermal stratification and adhesion to the basement membrane zone (BMZ) in skin. We used three assay systems, HFK adhesion to purified extracellular matrix (ECM) ligands and endogenous secreted ECM, localization of integrins in focal adhesions (FAs), and inhibition of HFK adhesion with mAbs to conclude: (a) A new anti-alpha 3 beta 1 mAb, P1F2, localized alpha 3 beta 1 in FAs on purified laminin greater than fibronectin/collagen, indicating that laminin was the best exogeneous ligand for alpha 3 beta 1. However, in long term culture, alpha 3 beta 1 preferentially codistributed in and around FAs with secreted laminin-containing ECM, in preference to exogenous laminin. Anti-alpha 3 beta 1, mAb P1B5, detached prolonged cultures of HFKs from culture plates or from partially purified HFK ECM indicating that interaction of alpha 3 beta 1 with the secreted laminin-containing ECM was primarily responsible for HFK adhesion in long term culture. (b) In FA assays, alpha 2 beta 1 localized in FAs conincident with initial HFK adhesion to exogenous collagen, but not laminin or fibronectin. However, in inhibition assays, anti-alpha 2 beta 1 inhibited initial HFK adhesion to both laminin and collagen. Thus, alpha 2 beta 1 contributes to initial HFK adhesion to laminin but alpha 3 beta 1 is primarily responsible for long-term HFK adhesion to secreted laminin-containing ECM. (c) Serum or Ca2(+)-induced aggregation of HFKs resulted in relocation of alpha 2 beta 1 and alpha 3 beta 1 from FAs to cell-cell contacts. Further, cell-cell adhesion was inhibited by anti-alpha 3 beta 1 (P1B5) and a new anti-beta 1 mAb (P4C10). Thus, interaction of alpha 3 beta 1 with either ECM or membrane coreceptors at cell-cell contacts may facilitate Ca2(+)-induced HFK aggregation. (d) It is suggested that interaction of alpha 3 beta 1 with a secreted, laminin-containing ECM in cultured HFKs, duplicates the role of alpha 3 beta 1 in basal cell adhesion to the BMZ in skin. Further, relocation of alpha 2 beta 1 and alpha 3 beta 1 to cell-cell contacts may result in detachment of cells from the BMZ and increased cell-cell adhesion in the suprabasal cells contributing to stratification of the skin.

Antibodies↗

Hepatocyte adhesion to carbohydrate-derivatized surfaces. II. Regulation of cytoskeletal organization and cell morphology.

Rat hepatic lectins mediate adhesion of isolated rat hepatocytes to synthetic surfaces derivatized with galactosides. Initial weak adhesion is followed by rapid adhesion strengthening. After hepatocytes contact galactose-derivatized gels, the hepatic lectins move rapidly into an inaccessible patch at the adhesive surface (Weisz, O. A., and R. L. Schnaar. 1991. J. Cell Biol. 115:485-493). Hepatic lectin patching, which occurs both at 37 degrees C and 4 degrees C, is not responsible for adhesion strengthening, which does not occur at 4 degrees C. Of various cytoskeletal and metabolic perturbants tested, only a combination of hyperosmotic medium, colchicine, and cytochalasin caused a marked (72%) reduction of adhesion strengthening (without reducing weak cell adhesion). Clathrin and actin were readily detected in the adhesive patch by immunofluorescence microscopy. Rat hepatocytes also adhered avidly to surfaces derivatized with asialofetuin, a high-affinity ligand for the rat hepatic lectins. However, hepatic lectin molecules did not migrate into a patch on the asialofetuin-derivatized surface, suggesting that hepatic lectin-asialofetuin binding may have resulted in the rapid formation of a ring of essentially irreversibly adherent receptors that prevented diffusion of additional lectin molecules into the contact site. The cells were unable to increase their adhesive contact area by flattening onto the derivatized surface. Treatment of cells with cytochalasin, however, did result in an increase in the size of the contact area. Cells adhering to surfaces derivatized with an adhesion-promoting peptide (containing an arg-gly-asp sequence) had larger contact areas than those adhering to galactoside-derivatized surfaces. A model is proposed in which carbohydrate-mediated adhesion causes specific reorganization of cytoskeletal components, leading to strengthened adhesion and a characteristic spherical cell morphology.

Actins↗

Thrombin adhesive properties: induction by plasmin and heparan sulfate.

We have previously demonstrated that chemically modified thrombin preparations induce endothelial cell (EC) adhesion, spreading and cytoskeletal reorganization via an Arg-Gly-Asp (RGD) sequence and the alpha v beta 3 integrin. Native thrombin, however, did not exhibit adhesive properties, consistent with crystal structure analysis, showing that Gly-Asp residues of the RGD epitope are buried within the molecule. We have now identified a possible physiological mean of converting thrombin to an adhesive protein. Plasmin, the major end product of the fibrinolytic system, converted thrombin to an adhesive protein for EC in a time and dose-dependent manner. EC adhesion and spreading was also induced by a low molecular weight (approximately 3,000 D) cleavage fragment generated upon incubation of thrombin with plasmin. Cell adhesion mediated by this fragment was completely inhibited by the synthetic peptide GRGDSP. Conversion of thrombin to an adhesive molecule was significantly enhanced in the presence of heparin or heparan sulfate, while other glycosaminoglycans (GAGs) (e.g., dermatan sulfate, keratan sulfate, chondroitin sulfate) had no effect. The role of cell surface heparan sulfate in thrombin conversion to EC adhesive protein was investigated using CHO cell mutants defective in various aspects of GAG synthesis. Incubation of both thrombin and a suboptimal amount of plasmin on the surface of formaldehyde fixed wild-type CHO-KI cells resulted in an efficient conversion of thrombin to an adhesive molecule, as indicated by subsequent induction of EC attachment. In contrast, there was no effect to incubation of thrombin and plasmin with fixed CHO mutant cells lacking both heparan sulfate and chondroitin sulfate, or with cells expressing no heparan sulfate and a three-fold increase in chondroitin sulfate. A similar gain of adhesive properties was obtained upon incubation of thrombin and plasmin in contact with native, but not heparinase-treated extracellular matrix (ECM) produced by cultured ECs. It appears that cell surface and ECM-associated heparan sulfate modulate thrombin adhesive properties through its heparin binding site in a manner that enables suboptimal amounts of plasmin to expose the RGD domain. Our results demonstrate, for the first time, a significant modulation of thrombin molecule by heparin, resulting in its conversion to a potent adhesive protein for ECs. This conversion is most effective in contact with cell surfaces, basement membranes and ECM.

Animals↗