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Black pigment gallstones are found with increased frequency compared with cholesterol gallstones within the rokytansky-aschoff sinuses of the gallbladder
Biliary Disorders A517
April 1998 G2106 A NEW BIOMATERIAL MORE RESISTENT TO INFECTION TO B E USED TO MANUFACTURE BILIARY ENDOPROSTHESES. Cetta F., *Oggiani M., **Rappuoli R., Montalto G., Zuckermaun M., **Magnani A., Gori M., Baldi C., *Valensin P.E., **Barbucci R. Institute of Surgical Clinics, *Microbiology and **Department of Chemical Science and Technology, University of Siena, Italy. Biliary endoprostheses invariably undergo obstruction because of infection. Therefore, they are the most suitable model to be used to test the advantages of new biomaterials in comparison with previous "raditional"materials in terms of increased resistence to infection. It has previously been shown that biliary endoprostheses obstruct in 100% of cases due to clog formation because of biliary infection (1-2). In particular, the material resulting in stent clogging has the same composition as brown pigment stones (calcium bilirubinate and palmitate, with low cholesterol) and the initial step in the occlusion of endoprostheses is the formation of a proteic biofilm, which adheres to the internal surface of the prosthesis. In order to prevent or delay the formation of the proteic biofilm, a new biomaterial has been deviced and then successfully manufactured, which was not a new antibacterial "coating" of the prosthesis inner surface, with all known problems (short duration of the antibacterial effect) and side-effects (long-term bioaetivity of Ag+ or other metal ions, including cytotoxicity, sensitization, mutagenicity, etc.). On the contrary, the new biomaterial is a polymer, which is "constitutively" resistent to the adhesion of the biofilm to the inner surface of the prosthesis, because of its physical-chemical properties. The new prosthesis (PUPA) contained HyalS3.5 (Hyaluronic acid at different grade of sulfatation) that was crosslinked directly to the poliurethane (PU) chains, which made the bulk of the endoprosthesis, through hexamethylendiisocyanate (HMDI). IR (Infra-red spectroscopy) analysis showed that HMDI molecules crosslinked both to PU and HyalS3.5 chains. Because of different hydrophilic characteristics of these two polymers (PU is hydrophobic, whereas HyalS3.5 is hydrophilic) the chains of the latter remained in the bulk material in dry state. On the contrary, hydrophilic chains of HyalS3,5 moved to the inner surface in water solution, so enabling themselves to play their biological properties. HyalS3,5 showed a significant inhibitory activity versus adhesion of thrombin and other proteic molecules with thrombogenic activity, because of its stable linkage with heparin and heparin-like molecules. Initial "in vitro" testing of the prototype with cultures of lithogenenic bacteria showed reduced adhesion of the proteic biofilm. If resistence to infection will be further confirmed "in vivo", in particular in humans, PUPA could be used to manufacture every type of biological devices, i.e. not only biliary endoprostheses, but also urinary catheters and urethral stents, vascular prosthesis, breast and knee prostheses and all biomedical devices, which are at risk of infection. [1] Cetta F. Hepatology 1986, 6:482-489; [2] Cetta F. Am J Gastroenterol 1997, 92:542-543. • G2107 BILE INFECTION PRECEDES, NOT FOLLOWS, THE FORMATION OF CLOGS OBSTRUCTING BILIARY ENDOPROSTHESES AND PLAYS A BASIC ROLE IN THE CLOGGING PROCESS. Cetta F., °Cariati A., Cetta D., Zuckermann M., Piro P., Baldi C., God M., Montalto G. Institute of Surgical Clinics, University of Siena; °Institute of Surgical Anatomy, University of Genoa; Italy. Twenty-one patients had treatment of non resectable tumors of the pancreas (n=ll), bile tract (n=5) or of the ampulla of Vater (n=5) by biliary endoprostheses (EPST) (10 French, outer diameter 3.2 mm, inner diameter 2.4 mm). Bile samples were obtained for bile culture before EPST placement. Culture was positive in 14 of the 21 patients (66%). EPST were removed within an average interval of 131 days (range 55-416 days) after implantation. Deposits obstructing the EPST were examined by stereomicroscopy and scanning electron microscopy and analyzed quantitatively by FTIR spectroscopy. All the deposits contained, in addition to small amounts of cholesterol, calcium palmitate and bilirubinate. In particular, calcium palmitate, which is a typical component of brown stones, was found both in the center and in periphery of the clogs, even if its concentration was significantly greater in the inner than in the outer portion (p=0,005) when compared with bilirubinate. At removal of EPST, bacteria were present in 100% of cases, usually showing a polymicrobic association (E. Coli, Pseudomonas, Proteus sp, Klebsiella). It is suggested that material resulting in stent clogging has the same composition as brown pigment stones (1) and then also the same pathogenesis (2). In particular, bile infection, as in brwon stone pathogenesis, precedes, not follows the clogging of the EPST, and is responsible for the formation of the brown deposits. Sphincterotomy and placement of a permanent foreign body within the bile tract are basic factors for bile contamination and growth of bacteria in patients with previous sterile bile. However, in addition to the diameter of the EPST and other technical parameters, factors related to the patient, as age and types of bacteria present in the duodenum, also play a role in stent clogging, which has to be considered a multifactorial process. [1] Cetta F. Hepatology 1986; 6:482-489. [2] Cetta F. Am. J. GastroenteroL 1997; 92:542-543.
G2108
BLACK PIGMENT GALLSTONES ARE FOUND WITH INCREASED FREQUENCY COMPARED WITH CHOLESTEROL GALLSTONES WITHIN THE ROKYTANSKY-ASCHOFF SINUSES OF THE GALLBLADDER. Cetta F., Lombardo F., *Cariati A., **Malet P.F., Baldi C., Montalto G., Piro P., Zuckermann M. Institute of Surgical Clinics, University of Siena; *Institute of Surgical Anatomy, University of Genoa; Italy. **Liver Unit, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas. Black pigment gallstones (BGS) contain mostly calcium salts of bilirubinate, carbonate and phosphate, and little to no cholesterol. Increased concentration of unconjugated bilimbi in the bile has been suggested as a major pathogenetic factor. BGS can be found not only within the gallbladder (GB) lumen, but also within the Rokytansky-Aschoff (R-A) sinuses, in patients with focal or diffuse adenomyomatosis (ADM). We have previously shown that BGS have been found within R-A sinuses, either as unique stones or in association with cholesterol (Ch) stones in the main GB lumen (1). The aim of the study was to determine the frequency of BGS within the R-A sinuses. Among 1421 consecutive patients with gallstones (GS), 137 patients (58 M; 79 W, mean age 61.4, range 6 to 90) had BGS as unique GS in the GB (9.6% of all gallbladder stones). In particular, 51 patients had macroscopically evident intraparietal microstones, either as unique stones or in association with similar or different types of GS in the main GB lumen. 47 of these GS were black, 4 were cholesterol. This ratio was significantly different from the relative incidence of Ch and black GS in the main lumen (1149 vs 137) (p<0.001). While 50 patients with BGS (36.5%) were associated with known risk factors, such as hemolysis, cirrhosis and old age, there were 35 patients (25.5%), who did not have any of the typical risk factors for BGS. All 35 patients had evidence of ADM or at least of an increase in the number and depth of the R-A sinuses of the GB wall. This is in contrast with the finding of ADM and R-A sinuses in only 25% of the 1149 patients with Ch GS (p<0.001). 27 of these 35 patients had intraparietal BGS, which were also intraluminal in 20 cases. CONCLUSIONS: Present data suggest that parietal factors, such as ADM, play a basic role in the formation of some BGS. R-A sinuses could play a role in the formation of GS different from those initially formed in the main lumen of the GB. R-A sinuses in patients with previous large Ch GS could be a consequence of the repeated intermittent occlusion of the infundibulum by the previous Ch GS, whereas congenital predisposition should be hypothesized in patients with R-A sinuses and BGS as unique stones. The reason why BGS form more frequently than Ch GS in microenvironments with local bile stasis, such as R-A sinuses, are not yet clear. [1] Cetta F. et al. Dig. Dis. Sci. 1995; 40:534-538. G2109 DOES HELICOBACTER PYLORI PLAY A ROLE IN THE PATHOGENESlS OF SOME TYPES OF GALLSTONES. Cetta F., Montalto G., Cetta D., Zuckermann M., Gori M., Baldi C., Piro P., *Figura N. Institute of Surgical Clinics and *Internal Medicine, University of Siena, Italy. BACKGROUND AND AIMS Gallstones (GS) are not a unique entity, but a heterogeneous disease, including cholesterol, mixed and pigment GS (black and brown). In particular, brown GS can be considered as a true infectious disease, mainly due to bile infection by E. Coli (1). Helicobacter Pylori (HP) has many similarities with E. Coli: (i) it produces enzymes responsible for hydrolysis of some constituents of the bile and calcium salt precipitation; (ii) determines chronic inflammation, possibly altering gallblader motility; (iii) produces Ig and other "nucleating proteins"; (iv) acting as a foreign body, can facilitate heterologous nucleation. The aim of the study has been to establish whether HP infection: (i) is associated with GS and in particular with which types of GS; (ii) could play a role in GS formation. METHODS 50 consecutive patients with different types of GS were analyzed for the presence of HP antibodies, antigens and/or genes in the bile obtained at operation. All patients had blood samples to detect HP infection in the serum. Western blotting and PCR were used to determine presence of antigens and genes. RESULTS 37 patients (74%) were seropositive for HP. HPB antibodies in the bile were only found in patients with HP antibodies in the blood. In particular, at least one of the markers of HP presence in the bile was found in 1 of 4 with black stones; 1 of 5 patients with solitary Ch stones; 2 of 7 with multiple spherical Ch GS; 5 of 7 with faceted stones; 9 Of 11 with composite GS, i.e. stones of different types within the same gallbladder; 3 of 3 with brown GS in the common duct (p<0.05). In particular, antigens were always found in patients with HP infection in the common duct bile after sphincterotomy. CONCLUSIONS Present data are preliminary and further confirmation is required in larger series. They suggest that HP presence in the bile is non homogeneously associated with the various types of GS. Even if HP certainly does not play a pathogenetic role similar to that of E.Coli in brown stone formation, the hypothesis that HP could be a co-factor for the formation of some subtypes of GS, maybe multiple Ch, mixed or composite GS, cannot be excluded. [1] Cetta F. Ann. Surg. 1991; 213:315-326.
April 1998 G2106 A NEW BIOMATERIAL MORE RESISTENT TO INFECTION TO B E USED TO MANUFACTURE BILIARY ENDOPROSTHESES. Cetta F., *Oggiani M., **Rappuoli R., Montalto G., Zuckermaun M., **Magnani A., Gori M., Baldi C., *Valensin P.E., **Barbucci R. Institute of Surgical Clinics, *Microbiology and **Department of Chemical Science and Technology, University of Siena, Italy. Biliary endoprostheses invariably undergo obstruction because of infection. Therefore, they are the most suitable model to be used to test the advantages of new biomaterials in comparison with previous "raditional"materials in terms of increased resistence to infection. It has previously been shown that biliary endoprostheses obstruct in 100% of cases due to clog formation because of biliary infection (1-2). In particular, the material resulting in stent clogging has the same composition as brown pigment stones (calcium bilirubinate and palmitate, with low cholesterol) and the initial step in the occlusion of endoprostheses is the formation of a proteic biofilm, which adheres to the internal surface of the prosthesis. In order to prevent or delay the formation of the proteic biofilm, a new biomaterial has been deviced and then successfully manufactured, which was not a new antibacterial "coating" of the prosthesis inner surface, with all known problems (short duration of the antibacterial effect) and side-effects (long-term bioaetivity of Ag+ or other metal ions, including cytotoxicity, sensitization, mutagenicity, etc.). On the contrary, the new biomaterial is a polymer, which is "constitutively" resistent to the adhesion of the biofilm to the inner surface of the prosthesis, because of its physical-chemical properties. The new prosthesis (PUPA) contained HyalS3.5 (Hyaluronic acid at different grade of sulfatation) that was crosslinked directly to the poliurethane (PU) chains, which made the bulk of the endoprosthesis, through hexamethylendiisocyanate (HMDI). IR (Infra-red spectroscopy) analysis showed that HMDI molecules crosslinked both to PU and HyalS3.5 chains. Because of different hydrophilic characteristics of these two polymers (PU is hydrophobic, whereas HyalS3.5 is hydrophilic) the chains of the latter remained in the bulk material in dry state. On the contrary, hydrophilic chains of HyalS3,5 moved to the inner surface in water solution, so enabling themselves to play their biological properties. HyalS3,5 showed a significant inhibitory activity versus adhesion of thrombin and other proteic molecules with thrombogenic activity, because of its stable linkage with heparin and heparin-like molecules. Initial "in vitro" testing of the prototype with cultures of lithogenenic bacteria showed reduced adhesion of the proteic biofilm. If resistence to infection will be further confirmed "in vivo", in particular in humans, PUPA could be used to manufacture every type of biological devices, i.e. not only biliary endoprostheses, but also urinary catheters and urethral stents, vascular prosthesis, breast and knee prostheses and all biomedical devices, which are at risk of infection. [1] Cetta F. Hepatology 1986, 6:482-489; [2] Cetta F. Am J Gastroenterol 1997, 92:542-543. • G2107 BILE INFECTION PRECEDES, NOT FOLLOWS, THE FORMATION OF CLOGS OBSTRUCTING BILIARY ENDOPROSTHESES AND PLAYS A BASIC ROLE IN THE CLOGGING PROCESS. Cetta F., °Cariati A., Cetta D., Zuckermann M., Piro P., Baldi C., God M., Montalto G. Institute of Surgical Clinics, University of Siena; °Institute of Surgical Anatomy, University of Genoa; Italy. Twenty-one patients had treatment of non resectable tumors of the pancreas (n=ll), bile tract (n=5) or of the ampulla of Vater (n=5) by biliary endoprostheses (EPST) (10 French, outer diameter 3.2 mm, inner diameter 2.4 mm). Bile samples were obtained for bile culture before EPST placement. Culture was positive in 14 of the 21 patients (66%). EPST were removed within an average interval of 131 days (range 55-416 days) after implantation. Deposits obstructing the EPST were examined by stereomicroscopy and scanning electron microscopy and analyzed quantitatively by FTIR spectroscopy. All the deposits contained, in addition to small amounts of cholesterol, calcium palmitate and bilirubinate. In particular, calcium palmitate, which is a typical component of brown stones, was found both in the center and in periphery of the clogs, even if its concentration was significantly greater in the inner than in the outer portion (p=0,005) when compared with bilirubinate. At removal of EPST, bacteria were present in 100% of cases, usually showing a polymicrobic association (E. Coli, Pseudomonas, Proteus sp, Klebsiella). It is suggested that material resulting in stent clogging has the same composition as brown pigment stones (1) and then also the same pathogenesis (2). In particular, bile infection, as in brwon stone pathogenesis, precedes, not follows the clogging of the EPST, and is responsible for the formation of the brown deposits. Sphincterotomy and placement of a permanent foreign body within the bile tract are basic factors for bile contamination and growth of bacteria in patients with previous sterile bile. However, in addition to the diameter of the EPST and other technical parameters, factors related to the patient, as age and types of bacteria present in the duodenum, also play a role in stent clogging, which has to be considered a multifactorial process. [1] Cetta F. Hepatology 1986; 6:482-489. [2] Cetta F. Am. J. GastroenteroL 1997; 92:542-543.
G2108
BLACK PIGMENT GALLSTONES ARE FOUND WITH INCREASED FREQUENCY COMPARED WITH CHOLESTEROL GALLSTONES WITHIN THE ROKYTANSKY-ASCHOFF SINUSES OF THE GALLBLADDER. Cetta F., Lombardo F., *Cariati A., **Malet P.F., Baldi C., Montalto G., Piro P., Zuckermann M. Institute of Surgical Clinics, University of Siena; *Institute of Surgical Anatomy, University of Genoa; Italy. **Liver Unit, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas. Black pigment gallstones (BGS) contain mostly calcium salts of bilirubinate, carbonate and phosphate, and little to no cholesterol. Increased concentration of unconjugated bilimbi in the bile has been suggested as a major pathogenetic factor. BGS can be found not only within the gallbladder (GB) lumen, but also within the Rokytansky-Aschoff (R-A) sinuses, in patients with focal or diffuse adenomyomatosis (ADM). We have previously shown that BGS have been found within R-A sinuses, either as unique stones or in association with cholesterol (Ch) stones in the main GB lumen (1). The aim of the study was to determine the frequency of BGS within the R-A sinuses. Among 1421 consecutive patients with gallstones (GS), 137 patients (58 M; 79 W, mean age 61.4, range 6 to 90) had BGS as unique GS in the GB (9.6% of all gallbladder stones). In particular, 51 patients had macroscopically evident intraparietal microstones, either as unique stones or in association with similar or different types of GS in the main GB lumen. 47 of these GS were black, 4 were cholesterol. This ratio was significantly different from the relative incidence of Ch and black GS in the main lumen (1149 vs 137) (p<0.001). While 50 patients with BGS (36.5%) were associated with known risk factors, such as hemolysis, cirrhosis and old age, there were 35 patients (25.5%), who did not have any of the typical risk factors for BGS. All 35 patients had evidence of ADM or at least of an increase in the number and depth of the R-A sinuses of the GB wall. This is in contrast with the finding of ADM and R-A sinuses in only 25% of the 1149 patients with Ch GS (p<0.001). 27 of these 35 patients had intraparietal BGS, which were also intraluminal in 20 cases. CONCLUSIONS: Present data suggest that parietal factors, such as ADM, play a basic role in the formation of some BGS. R-A sinuses could play a role in the formation of GS different from those initially formed in the main lumen of the GB. R-A sinuses in patients with previous large Ch GS could be a consequence of the repeated intermittent occlusion of the infundibulum by the previous Ch GS, whereas congenital predisposition should be hypothesized in patients with R-A sinuses and BGS as unique stones. The reason why BGS form more frequently than Ch GS in microenvironments with local bile stasis, such as R-A sinuses, are not yet clear. [1] Cetta F. et al. Dig. Dis. Sci. 1995; 40:534-538. G2109 DOES HELICOBACTER PYLORI PLAY A ROLE IN THE PATHOGENESlS OF SOME TYPES OF GALLSTONES. Cetta F., Montalto G., Cetta D., Zuckermann M., Gori M., Baldi C., Piro P., *Figura N. Institute of Surgical Clinics and *Internal Medicine, University of Siena, Italy. BACKGROUND AND AIMS Gallstones (GS) are not a unique entity, but a heterogeneous disease, including cholesterol, mixed and pigment GS (black and brown). In particular, brown GS can be considered as a true infectious disease, mainly due to bile infection by E. Coli (1). Helicobacter Pylori (HP) has many similarities with E. Coli: (i) it produces enzymes responsible for hydrolysis of some constituents of the bile and calcium salt precipitation; (ii) determines chronic inflammation, possibly altering gallblader motility; (iii) produces Ig and other "nucleating proteins"; (iv) acting as a foreign body, can facilitate heterologous nucleation. The aim of the study has been to establish whether HP infection: (i) is associated with GS and in particular with which types of GS; (ii) could play a role in GS formation. METHODS 50 consecutive patients with different types of GS were analyzed for the presence of HP antibodies, antigens and/or genes in the bile obtained at operation. All patients had blood samples to detect HP infection in the serum. Western blotting and PCR were used to determine presence of antigens and genes. RESULTS 37 patients (74%) were seropositive for HP. HPB antibodies in the bile were only found in patients with HP antibodies in the blood. In particular, at least one of the markers of HP presence in the bile was found in 1 of 4 with black stones; 1 of 5 patients with solitary Ch stones; 2 of 7 with multiple spherical Ch GS; 5 of 7 with faceted stones; 9 Of 11 with composite GS, i.e. stones of different types within the same gallbladder; 3 of 3 with brown GS in the common duct (p<0.05). In particular, antigens were always found in patients with HP infection in the common duct bile after sphincterotomy. CONCLUSIONS Present data are preliminary and further confirmation is required in larger series. They suggest that HP presence in the bile is non homogeneously associated with the various types of GS. Even if HP certainly does not play a pathogenetic role similar to that of E.Coli in brown stone formation, the hypothesis that HP could be a co-factor for the formation of some subtypes of GS, maybe multiple Ch, mixed or composite GS, cannot be excluded. [1] Cetta F. Ann. Surg. 1991; 213:315-326.