Experimental Liver Cirrhosis Inhibits Restenosis after Balloon Angioplasty.

The effect of liver cirrhosis on vascular remodeling in vivo remains unknown. Therefore, this study investigates the influence of cholestatic liver cirrhosis on carotid arterial remodeling. A total of 79 male Sprague Dawley rats underwent bile duct ligation (cirrhotic group) or sham surgery (control group) and 28 days later left carotid artery balloon dilatation; 3, 7, 14 and 28 days after balloon dilatation, the rats were euthanized and carotid arteries were harvested. Histological sections were planimetrized, cell counts determined, and systemic inflammatory parameters measured. Up to day 14 after balloon dilatation, both groups showed a comparable increase in neointima area and degree of stenosis. By day 28, however, both values were significantly lower in the cirrhotic group (% stenosis: 20 ± 8 vs. 42 ± 10, p = 0.010; neointimal area [mm2]: 0.064 ± 0.025 vs. 0.138 ± 0.025, p = 0.024). Simultaneously, cell density in the neointima (p = 0.034) and inflammatory parameters were significantly higher in cirrhotic rats. This study demonstrates that cholestatic liver cirrhosis in rats substantially increases neointimal cell consolidation between days 14 and 28. Thereby, consolidation proved important for the degree of stenosis. This may suggest that patients with cholestatic cirrhosis are at lower risk for restenosis after coronary intervention.

Cirrhotic Cardiomyopathy Following Bile Duct Ligation in Rats-A Matter of Time?

Cirrhotic patients often suffer from cirrhotic cardiomyopathy (CCM). Previous animal models of CCM were inconsistent concerning the time and mechanism of injury; thus, the temporal dynamics and cardiac vulnerability were studied in more detail. Rats underwent bile duct ligation (BDL) and a second surgery 28 days later. Cardiac function was assessed by conductance catheter and echocardiography. Histology, gene expression, and serum parameters were analyzed. A chronotropic incompetence (Pd31 < 0.001) and impaired contractility at rest and a reduced contractile reserve (Pd31 = 0.03, Pdob-d31 < 0.001) were seen 31 days after BDL with increased creatine (Pd35, Pd42, and Pd56 < 0.05) and transaminases (Pd31 < 0.001). A total of 56 days after BDL, myocardial fibrosis was seen (Pd56 < 0.001) accompanied by macrophage infiltration (CD68: Pgroup < 0.001) and systemic inflammation (TNFα: Pgroup < 0.001, white blood cell count: Pgroup < 0.001). Myocardial expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) was increased after 31 (Pd31 < 0.001) and decreased after 42 (Pd42 < 0.001) and 56 days (Pd56 < 0.001). Caspase-3 expression was increased 31 and 56 days after BDL (Pd31 = 0.005; Pd56 = 0.005). Structural changes in the myocardium were seen after 8 weeks. After the second surgery (second hit), transient myocardial insufficiency with secondary organ dysfunction was seen, characterized by reduced contractility and contractile reserve.

A Proof-of-Concept Preclinical Study Using a Novel Thermal Insulation Device in a Porcine Kidney Auto-Transplantation Model.

Ischemia-reperfusion injury remains a fundamental problem during organ transplantation logistics. One key technical factor is the rapid allograft rewarming during the time of vascular reconstruction in the recipient. In this pilot study, a new thermal insulation bag (TIB) for organ transplantation was used. Insulation capacity, tissue compatibility, and usability were tested initially ex vivo on porcine kidneys (n = 24) followed by the first in vivo usage. Fourteen female German landrace pigs underwent kidney auto-transplantation after 24 h cold storage (4 °C). During the implantation process the kidney was either insulated with the new TIB, or it was not thermo-protected at all, which represents the clinical standard. In this proof-of-concept study, the usability (knife-to-skin-time) and the general thermal capacity (30 min warm storage at 38 °C ex vivo p < 0.001) was shown. The clinical outcome showed significant differences in the determination of CRP and pi-GST levels. Syndecan-1 Antibody staining showed clear significant higher counts in the control group (p < 0.01) indicating epithelial damage. However, the effect on renal outcomes in not severely pre-damaged kidneys does not appear to be conclusively significant. A close follow-up study is warranted, especially in the context of marginal organs or in cases where anastomosis-times are prolonged due to surgical complexity (e.g., multiple vessels and complex reconstructions).

The Impact of a Nitric Oxide Synthase Inhibitor (L-NAME) on Ischemia⁻Reperfusion Injury of Cholestatic Livers by Pringle Maneuver and Liver Resection after Bile Duct Ligation in Rats.

The Pringle maneuver (PM) has been widely used to control blood loss during liver resection. However, hepatic inflow occlusion can also result in hepatic ischemia-reperfusion injury (IRI), especially in patients with a cholestatic, fibrotic, or cirrhotic liver. Here we investigate a nitric oxide synthase (NOS) inhibitor N-Nitroarginine methyl ester (L-NAME) on IRI after the PM and partial hepatectomy of cholestatic livers induced by bile duct ligation (BDL) in rats. Control group (non-BDL/no treatment), BDL + T group (BDL/L-NAME treatment) and BDL group (BDL/no treatment) were analyzed. Cholestasis was induced by BDL in the L-NAME and BDL group and a 50% partial hepatectomy with PM was performed. L-NAME was injected before PM in the BDL + T group. Hepatocellular damage, portal venous flow, microcirculation, endothelial lining, and eNOS, iNOS, interleukin (IL)-6, and transforming growth factor-ß (TGF-ß) were evaluated. Microcirculation of the liver in the BDL + T group tended to be higher. Liver damage and apoptotic index were significantly lower and Ki-67 labeling index was higher in the BDL + T group while iNOS and TGF-ß expression was decreased. This was corroborated by a better preserved endothelial lining. L-NAME attenuated IRI following PM and improved proliferation/regeneration of cholestatic livers. These positive effects were considered as the result of improved hepatic microcirculation, prevention of iNOS formation, and TGF-ß mRNA upregulation.

AK03, a new recombinant fibrinogenase prevents abdominal adhesions in a rat model without systemic side effects.

BACKGROUND: Abdominal adhesions are one of the most common complications after abdominal surgery, and fibrin is suspected to be a crucial component. The aim of the current study was an in vivo evaluation of a new recombinant fibrinogenase (AK03) in two animal models. METHODS: Sixty-four rats were randomly divided into four groups (sodium chloride [NaCl], icodextrin, AK03 low dose, and AK03 high dose) and evaluated at two time endpoints. Adhesion model comprised both a visceral defect (terminal ileum) and parietal defect. Test (AK03) and control substances (NaCl and icodextrin) were administered intraperitoneally after setting the intraabdominal defects. A second dose was administered 24 h after surgery. Plasma fibrinogen values were taken at baseline and after 7 and 21 d, respectively. Rats were sacrificed after 7 or 21 d for macroscopic (Diamond score) and immunohistochemical investigations. RESULTS: After 7 and 21 d, the Diamond score of postsurgical adhesions were significantly lower in both AK03-treated groups compared with NaCl control group (P = 0.02). There were no unspecific systemic side effects in both treatment groups and no decrease in plasma fibrinogen concentration. In none of the four groups was there any evidence for impaired wound repair. Microscopically in the area of the parietal defect, we saw less cluster of differentiation 3+ T-lymphocytes and cluster of differentiation 68+ macrophages in both groups receiving AK03 compared with the NaCl and icodextrin control groups. CONCLUSIONS: The results of this study indicate that the new recombinant fibrinogenase AK03 effectively prevents peritoneal adhesions without causing side effects, notably systemic fibrinogen depletion, bleeding, or impaired wound repair. Due to these results, future clinical studies may be promising.

In vivo imaging of antioxidant response element activity during liver regeneration after partial hepatectomy.

BACKGROUND: The nuclear factor-erythroid 2-related factor 2 (Nrf2) -antioxidant response element (ARE) pathway is important for the regulation of antioxidative stress response and detoxification. To activate the expression of its target genes, such as heme oxygenase-1 (HO-1) and NAD(P)H dehydrogenase (quinone) 1 (NQO1), Nrf2 binds to the ARE within the promoter region of these genes. Partial hepatectomy and consecutive liver regeneration lead to oxidative stress with activation of the Nrf2-ARE pathway. The aim of this study was to investigate ARE activity in vivo during liver regeneration after partial hepatectomy. MATERIALS AND METHODS: Transgenic ARE-luc mice were used. In these mice, the luciferase reporter gene is under the control of an ARE promoter element. Following 2/3 partial hepatectomy (PHx), mice underwent in vivo bioluminescence imaging up until the ninth postoperative day. In addition, liver tissue was analyzed by immunohistochemistry (Nrf2 and HO-1), quantitative reverse transcription-PCR (HO-1 and NQO1) and in vitro luminescence assays. RESULTS: Bioluminescence imaging revealed a significant increase in Nrf2-ARE activity after PHx. The signal maximum was recorded on the third day after PHx. Seven days postoperatively, the signal almost reached baseline levels. In immunohistochemistry, significantly more hepatocytes were positive for Nrf2 and HO-1 on the third postoperative day compared with baseline levels. The mRNA expression of HO-1 and NQO1 were significantly increased on day 3 as measured by qRT-PCR. CONCLUSIONS: This study demonstrated the time-dependent activation of the Nrf2-ARE system during liver regeneration in vivo. The transgenic ARE-luc mouse provided a convenient model for studying Nrf2-mediated gene expression noninvasively and may facilitate further experiments with therapeutic modulation of the antioxidative stress response.

Vascular anatomy of the small intestine-a comparative anatomic study on humans and pigs.

BACKGROUND: Porcine models are well established for studying intestinal anastomotic healing. In this study, we aimed to clarify the anatomic differences between human and porcine small intestines. Additionally, we investigated the influences of longitudinal and circular sutures on human small intestine perfusion. METHODS: Intestines were obtained from human cadavers (n = 8; small intestine, n = 51) and from pigs (n = 10; small intestine, n = 60). Vascularization was visualized with mennige gelatin perfusion and high-resolution mammography. Endothelial cell density was analyzed with immunohistochemistry and factor VIII antibodies. We also investigated the influence of suture techniques (circular anastomoses, n = 19; longitudinal sutures, n = 15) on vascular perfusion. RESULTS: Only human samples showed branching of mesenteric vessels. Compared to the pig, human vessels showed closer connections at the entrance to the bowel wall (p = 0.045) and higher numbers of intramural anastomoses (p < 0.001). Porcine main vessels formed in multifilament-like vessel bundles and displayed few intramural vessel anastomoses. Circular anastomoses induced a circular perfusion defect at the bowel wall; longitudinal anastomoses induced significantly smaller perfusion defects (p < 0.001). Both species showed higher vascular density in the jejunum than in the ileum (p < 0.001). Human samples showed similar vascular density within the jejunum (p = 0.583) and higher density in the ileum (p < 0.001) compared to pig samples. CONCLUSION: The results showed significant differences between human and porcine intestines. The porcine model remains the standard for studies on anastomotic healing because it is currently the only viable model for studying anastomosis and wound healing. Nevertheless, scientific interpretations must consider the anatomic differences between humans and porcine intestines.

Polyvinylidene Fluoride as a Suture Material: Evaluation of Comet Tail-Like Infiltrate and Foreign Body Granuloma.

BACKGROUND: Biocompatibility and tissue integration of a surgical suture are decisive factors for wound healing and therefore for the success of sutures. The optimal suture material is still under discussion. Polyvinylidene fluoride (PVDF) is described to have superior properties of biocompatibility and is therefore frequently used as a mesh component. Only little information is available about its use as a suture material. The aim of this study was to evaluate the biocompatibility of PVDF as a suture material in comparison to 5 different established sutures in a rat model. METHODS: In 30 male rats, a monofilamental PVDF suture (Resopren®) and 5 established control suture materials [polyester (Miralene®), polytetrafluoroethylene (Gore®), poliglecaprone (Monocryl®), polydioxanone (Monoplus®), polyglactin 910 (Vicryl®), USP size 3-0] were placed in the subcutaneous layer of the abdominal wall without knot or tension. After 3, 7 or 21 days, the abdominal walls were explanted for histopathological and immunohistochemical investigation with special regard to the size and quality of foreign body granuloma and the length of the comet tail-like infiltrate (CTI). RESULTS: The PVDF sutures showed the smallest size of foreign body granuloma (60 ± 14 µm) and the smallest CTI length (343 ± 60 µm) of all polymers after 21 days. Only PVDF (Resopren) and polydioxanone (Monoplus) showed a significant collagen I/III ratio increase between days 3 and 21 (p = 0.009 and p = 0.016). The quality of foreign body reaction regarding inflammation, proliferation and fibrotic remodeling was similar between all suture materials. CONCLUSIONS: Our data indicate that monofilamental PVDF sutures show a favorable foreign body reaction with small granuloma sizes and CTI length in comparison to established sutures. Its use as a suture material in general surgery could therefore be extended in the future. To reinforce these findings, further clinical studies need to be conducted.

Role of preferential cyclooxygenase-2 inhibition by meloxicam in ischemia/reperfusion injury of the rat liver.

BACKGROUND: Ischemia/reperfusion injury (IRI) is one of the major clinical problems in liver and transplant surgery. Livers subjected to warm ischemia in vivo often show a severe dysfunction and the release of numerous inflammatory cytokines and arachidonic acid metabolites. Cyclooxygenase (COX)-2 is the inducible isoform of an intracellular enzyme that converts arachidonic acid into prostaglandins. The aim of the study was to evaluate the effect of COX-2 inhibition and the role of Kupffer cells in IRI of the liver. METHODS: Male Wistar rats [250- 280 g body weight (BW)] were anesthetized and subjected to 30-min warm ischemia of the liver (Pringle's maneuver) and 60-min reperfusion after median laparotomy. The I/R group received no additional treatment. In the COX-2 inhibitor (COX-2I) group, the animals received 1 mg/kg BW meloxicam prior to operation. Gadolinium chloride (GdCl3) (10 mg/kg BW) was given 24 h prior to operation in the GdCl3 and GdCl3 + COX-2I groups for the selective depletion of Kupffer cells. The GdCl3 + COX-2I group received both GdCl3 and meloxicam treatment prior to operation. Blood and liver samples were obtained at the end of the experiments for further investigations. RESULTS: After 30 min of warm ischemia in vivo, severe hepatocellular damage was observed in the I/R group. These impairments could be significantly prevented by the selective COX-2 inhibition and the depletion of Kupffer cells. Alanine aminotransferase was significantly reduced upon meloxicam and GdCl3 treatment compared to the I/R group: I/R, 3,240 ± 1,262 U/l versus COX-2I, 973 ± 649 U/l, p < 0.001; I/R versus GdCl3, 1,611 ± 600 U/l, p < 0.05, and I/R versus GdCl3 + COX-2I, 1,511 ± 575 U/l, p < 0.01. Plasma levels of tumor necrosis factor alpha (TNF-α) were significantly reduced in the COX-2I treatment group compared to I/R (3.5 ± 1.5 vs. 16.3 ± 11.7 pg/ml, respectively; p < 0.05). Similarly, the amount of TxB2, a marker for COX-2 metabolism, was significantly reduced in the meloxicam treatment groups compared to the I/R group: I/R, 22,500 ± 5,210 pg/ml versus COX-2I, 1,822 ± 938 pg/ml, p < 0.001, and I/R versus GdCl3 + COX-2I, 1,530 ± 907 pg/ml, p < 0.001. All values are given as mean ± SD (n = 6). CONCLUSION: These results suggest that the inhibition of COX-2 suppressed the initiation of an inflammatory cascade by attenuating the release of TNF-α, which is an initiator of the inflammatory reaction in hepatic IRI. Therefore, we conclude that preferential inhibition of COX-2 is a possible therapeutic approach against warm IRI of the liver.

Coronary Microvascular Dysfunction in Acute Cholestasis-Induced Liver Injury.

BACKGROUND: Previous studies have shown cardiac abnormalities in acute liver injury, suggesting a potential role in the associated high mortality. METHODS: We designed an experimental study exploring the short-term effects of acute cholestasis-induced liver injury on cardiac function and structure in a rodent bile duct ligation (BDL) model to elucidate the potential interplay. Thirty-seven male Sprague-Dawley rats were subjected to BDL surgery (n = 28) or served as sham-operated (n = 9) controls. Transthoracic echocardiography, Doppler evaluation of the left anterior descending coronary artery, and myocardial contrast echocardiography were performed at rest and during adenosine and dobutamine stress 5 days after BDL. Immunohistochemical staining of myocardial tissue samples for hypoxia and inflammation as well as serum analysis were performed. RESULTS: BDL animals exhibited acute liver injury with elevated transaminases, bilirubin, and total circulating bile acids (TBA) 5 days after BDL (TBA control: 0.81 ± 2.54 µmol/L vs. BDL: 127.52 ± 57.03 µmol/L; p < 0.001). Concurrently, cardiac function was significantly impaired, characterized by reduced cardiac output (CO) and global longitudinal strain (GLS) in the echocardiography at rest and under pharmacological stress (CO rest control: 120.6 ± 24.3 mL/min vs. BDL 102.5 ± 16.6 mL/min, p = 0.041; GLS rest control: -24.05 ± 3.8% vs. BDL: -18.5 ± 5.1%, p = 0.01). Myocardial perfusion analysis revealed a reduced myocardial blood flow at rest and a decreased coronary flow velocity reserve (CFVR) under dobutamine stress in the BDL animals (CFVR control: 2.1 ± 0.6 vs. BDL: 1.7 ± 0.5 p = 0.047). Immunofluorescence staining indicated myocardial hypoxia and increased neutrophil infiltration. CONCLUSIONS: In summary, acute cholestasis-induced liver injury can lead to impaired cardiac function mediated by coronary microvascular dysfunction, suggesting that major adverse cardiac events may contribute to the mortality of acute liver failure. This may be due to endothelial dysfunction and direct bile acid signaling.

'The ideal mesh?'.

Currently, more than 200 different textile constructions, so-called 'meshes', are available for use world-wide in the more than 20 million operations performed annually for the reinforcement of tissues. As any reintervention at the mesh-tissue compound is a surgical challenge, sometimes resulting in almost untreatable defects, huge efforts are being made to improve the biological and functional performance of the meshes. Based on numerous experimental and clinical studies in the past 20 years, our understanding of them has improved markedly. This includes the biomechanical aspects and the histopathological evaluation of the recipient tissue. Sufficiently large pores as well as structural stability in case of mechanical strain have been identified to be crucial to reduce excessive inflammation and fibrosis. Furthermore, large pores prevent bridging of the foreign body reaction through the pore and thereby help to reduce clinical adverse events as erosion, shrinkage or pain. However, with regard to the many different indications for meshes, there will never be one single ideal mesh for all purposes. To achieve an optimal performance, every construction should be designed according to the specific functional requirements, charging the surgeon to identify the best mesh for his purpose.

Influence of 4% icodextrin solution on peritoneal tissue response and adhesion formation.

BACKGROUND: Postoperative peritoneal adhesion formation following abdominal surgery remains a relevant surgical problem. The application of soluble physico-chemical barriers like 4% icodextrin is one approach to protect the peritoneal surface from getting linked to adhesive scar. The aim of this study was to investigate the influence of 4% icodextrin on peritoneal tissue response both of visceral and parietal peritoneum, adhesion formation and wound healing. METHODS: 40 rats were divided into two groups. After creation of an intraabdominal defect, either 4% icodextrin (Adept®) or sodium chloride was applied. Animals were sacrificed after 7 and 21 days. Adhesions were scored by an adhesion score. Furthermore, immunohistochemical investigations were conducted to determine the discrete influence of icodextrin on the parietal and visceral peritoneal tissue responses (CD68+ macrophages, CD3+ T-lymphocytes, vimentin for mesenchymal cells, HBME-1 for mesothelial cells, and as components of wound healing COX-2, C-myc, catenin). RESULTS: Postoperative peritoneal adhesions were predominantly present in the sodium chloride group as compared to the icodextrin group (14/19 (74%) vs. 9/19 (47%); p = 0.048). The adhesion score however did not reveal any significant differences, (p = 0.614). Furthermore, the expression of vimentin in both the parietal and visceral peritoneum after 21 days was significantly lower in the icodextrin group than in the sodium chloride group (p = 0.038 and p = 0.028, respectively). No significant differences were observed for macrophages, lymphocytes, reperitonealisation or the expression of COX-2, C-myc or Catenin. CONCLUSIONS: The intraperitoneal application of 4% icodextrin reduces adhesion formation in comparison to sodium chloride. 4% icodextrin solution reduces the inflammatory and mesenchymal infiltrate in the wounded area, thus improving the ratio of mesothel cells to mesenchymal infiltrate. As demonstrated, icodextrin is able to ameliorate the local tissue response. Further experimental studies would be done to elaborate the impact on the early response of the adaptive immune system, which may then trigger the subsequent wound healing and tissue repair.

Influence of polypropylene mesh degradation on tissue inflammatory reaction.

Polypropylene degradation in vivo appears as mesh surface cracking and peeling. This aging process of the mesh, resulting in the lack of bio-stability, contradicts the requirement of biocompatibility. However, to date, it is still not clearly established how much this mesh degradation influences the local tissue response with subsequent clinical consequences. This study aims to find out whether mesh degradation is correlated with elevated inflammatory tissue reaction through analyzing 100 human PP meshes explanted from the pelvic floor. A degradation classification method, based on standard pathological H&E stained slides of the explanted mesh via light microscope, was developed to classify the mesh degradation into four classes (no, mild, moderate and severe degradation). The peri-filamentary tissue inflammatory reaction was analyzed by scoring the expression of the most common cell markers for the innate immune reaction: CD68 as marker for macrophage, CD86 for M1 subtype, CD163 for M2 subtype, CD3 for T-lymphocyte and CD15 for neutrophil granulocytes. The correlation between immune cell expression, degradation classification and time of implantation of the meshes are evaluated with Spearman-Rho-Test. Mesh degradation worsens significantly (p < .001) with longer time of implantation. The increasing tendency of CD68 expression by mesh with higher degradation class indicates that the number of macrophages increases with worsening mesh degradation. The significantly increased expression of CD163 and CD3 cell by severely degraded mesh demonstrate the increased number of M2 and T-Lymphocyte when mesh degradation becomes severe. None of the inflammatory cells show the usual declining expression with longer time of implantation. The result of this study suggests that the degradation of PP mesh results in an elevated local inflammatory reaction in female pelvic floor. A material with better bio-stability for mesh implant in pelvic floor is required.

A Deep-Learning-Computed Cancer Score for the Identification of Human Hepatocellular Carcinoma Area Based on a Six-Colour Multiplex Immunofluorescence Panel.

Liver cancer is one of the most frequently diagnosed and fatal cancers worldwide, with hepatocellular carcinoma (HCC) being the most common primary liver cancer. Hundreds of studies involving thousands of patients have now been analysed across different cancer types, including HCC, regarding the effects of immune infiltrates on the prognosis of cancer patients. However, for these analyses, an unambiguous delineation of the cancer area is paramount, which is difficult due to the strong heterogeneity and considerable inter-operator variability induced by qualitative visual assessment and manual assignment. Nowadays, however, multiplex analyses allow the simultaneous evaluation of multiple protein markers, which, in conjunction with recent machine learning approaches, may offer great potential for the objective, enhanced identification of cancer areas with further in situ analysis of prognostic immune parameters. In this study, we, therefore, used an exemplary five-marker multiplex immunofluorescence panel of commonly studied markers for prognosis (CD3 T, CD4 T helper, CD8 cytotoxic T, FoxP3 regulatory T, and PD-L1) and DAPI to assess which analytical approach is best suited to combine morphological and immunohistochemical data into a cancer score to identify the cancer area that best matches an independent pathologist's assignment. For each cell, a total of 68 individual cell features were determined, which were used as input for 4 different approaches for computing a cancer score: a correlation-based selection of individual cell features, a MANOVA-based selection of features, a multilayer perceptron, and a convolutional neural network (a U-net). Accuracy was used to evaluate performance. With a mean accuracy of 75%, the U-net was best capable of identifying the cancer area. Although individual cell features showed a strong heterogeneity between patients, the spatial representations obtained with the computed cancer scores delineate HCC well from non-cancer liver tissues. Future analyses with larger sample sizes will help to improve the model and enable direct, in-depth investigations of prognostic parameters, ultimately enabling precision medicine.

Choice of Polymer, but Not Mesh Structure Variation, Reduces the Risk of Bacterial Infection with Staphylococcus aureus In Vivo.

BACKGROUND: Synthetic mesh material is of great importance for surgical incisional hernia repair. The physical and biochemical characteristics of the mesh influence mechanical stability and the foreign body tissue reaction. The influence on bacterial infections, however, remains ill-defined. The aim of the present study was to evaluate the influence of a modified mesh structure with variation in filament linking on the occurrence of bacterial infection that is indicated by the occurrence of CD68+, CD4+, and CD8+ cells in two different materials. METHODS: A total of 56 male Sprague Dawley rats received a surgical mesh implant in a subcutaneous abdominal position. The mesh of two different polymers (polypropylene (PP) and polyvinylidenfluoride (PVDF)) and two different structures (standard structure and bold structure with higher filament linking) were compared. During the implantation, the meshes were infected with Staphylococcus (S.) aureus. After 7 and 21 days, meshes were explanted, and the early and late tissue responses to infection were histologically evaluated. RESULTS: Overall, the inflammatory tissue response was higher at 7 days when compared to 21 days. At 7 days, PP meshes of the standard structure (PP-S) showed the strongest inflammatory tissue response in comparison to all the other groups. At 21 days, no statistically significant difference between different meshes was detected. CD8+ cytotoxic T cells showed a significant difference at 21 days but not at 7 days. PP meshes of both structures showed a higher infiltration of CD8+ T cells than PVDF meshes. CD4+ T helper cells differed at 7 days but not at 21 days, and PVDF meshes in a bold structure showed the highest CD4+ T cell count. The number of CD68+ macrophages was also significantly higher in PP meshes in a standard structure when compared to PVDF meshes at 21 days. CONCLUSION: The inflammatory tissue response to S. aureus infection appears to be highest during the early period after mesh implantation. PP meshes showed a higher inflammatory response than PVDF meshes. The mesh material appears to be more important for the risk of infection than the variation in filament linking.

Reduction of activated macrophages after ischaemia-reperfusion injury diminishes oxidative stress and ameliorates renal damage.

BACKGROUND: Macrophages are major effectors of the local inflammatory response syndrome (LIRS) and influence the extent of ischaemia/reperfusion injury, thereby impacting organ function. Several subgroups of macrophages exist, representing distinct modes of action. The specific role of the subset expressing Fc gamma receptor (FcγR) 1 in the activated state of macrophages is poorly defined. METHODS: We induced a LIRS via 30 min of ischaemia in uninephrectomized rats, transgenic for the human FcγR1. Six hours after reperfusion, the treatment group was injected with a recombinant immunotoxin (IT) H22(scFv)-ETA' targeted against human FcγR1, which induced apoptosis of target cells. The placebo group received normal saline (NS). Contralateral kidneys served as healthy controls (Ctr). After 24 h of reperfusion, the animals were analysed. RESULTS: Targeted treatment with IT resulted in preserved renal function [NS versus IT treatment and baseline (creatinine: 69.2 ± 2.6, 54.7 ± 3.4 and 27.3 ± 1.0 µmol/L; P < 0.001)]. The number of all infiltrating monocytes were significantly reduced (CD68-positive cells per view field: NS 3.8 ± 0.4, IT 2.5 ± 0.2 and Ctr 1.2 ± 0.4; P < 0.05), renal histology improved and there was a reduced expression of renal fibronectin (NS 4.0 ± 0.4, IT 2.3 ± 0.2 and Ctr 1.1 ± 0.1; P < 0.001). Following IT administration, we also observed less expression of renal monocyte chemoattractant protein-1-positive cells per view field (NS 19.0 ± 1, IT 10.1 ± 0.8 and Ctr 2.0 ± 0.3; P < 0.001) as well as reduced systemic and local oxidative stress [serum malondialdehyde (MDA): NS 340 ± 30, IT 224 ± 36 versus baseline 140 ± 5 nmol/mL; P < 0.01]; renal MDA arbitrary units of fluorescence intensity: NS 3.7 ± 0.2, IT 1.8 ± 0.3 and Ctr 0.4 ± 0.2; P < 0.001. CONCLUSIONS: Reduction of FcγR1-up-regulated monocytic cells leads to preserved renal function and morphology in a rat model of ischaemia-triggered LIRS. Our results show that targeting activated macrophages is a valuable approach for ameliorating ischaemia-induced tissue injury.

In vivo MRI visualization of mesh shrinkage using surgical implants loaded with superparamagnetic iron oxides.

BACKGROUND: Prosthetic mesh implants are widely used in hernia surgery. To show long-term mesh-related complications such as shrinkage or adhesions, a precise visualization of meshes and their vicinity in vivo is important. By supplementing mesh fibers with ferro particles, magnetic resonance imaging (MRI) can help to delineate the mesh itself. This study aimed to demonstrate and quantify time-dependent mesh shrinkage in vivo by MRI. METHODS: Polyvinylidenfluoride (PVDF) meshes with incorporated superparamagnetic iron oxides (SPIOs) were implanted as an abdominal wall replacement in 30 rats. On days 1, 7, 14, or 21, MRI was performed using a gradient echo sequence with repetition time (TR)/echo time (TE) of 50/4.6 and a flip angle of 20°. The length, width, and area of the device were measured on axial, coronal, and sagittal images, and geometric deformations were assessed by surgical explantation. RESULTS: In all cases, the meshes were visualized and their area estimated by measuring the length and width of the mesh. The MRI presented a mean area shrinkage in vivo of 13% on day 7, 23% on day 14, and 23% on day 21. Postmortem measurements differed statistically from MRI, with a mean area shrinkage of 23% on day 7, 28% on day 14, and 30% on day 21. Ex vivo measurements of shrinkage showed in vivo measurements to be overestimated approximately 8%. Delineation of the mesh helped to show folding or adhesions close to the intestine. CONCLUSION: Loading of surgical meshes with SPIOs allows their precise visualization during MRI and guarantees an accurate in vivo assessment of their shrinkage. The authors' observation clearly indicates that shrinkage in vivo is remarkably less than that shown by illustrated explantation measurements. The use of MRI with such meshes could be a reliable technique for checking on proper operation of implanted meshes and showing related complications, obviating the need for exploratory open surgical revision.

Mesh biocompatibility: effects of cellular inflammation and tissue remodelling.

Mesh biocompatibility is basically determined by the foreign body reaction (FBR). In contrast to physiological wound healing and scar formation, the FBR at the host-tissue/biomaterial interface is present for the lifetime of the medical device. The cellular interactions at the mesh/tissue interface proceed over time ending up in a chronic inflammatory process. The time course of the FBR has been studied extensively and consists of three crucial steps that are protein absorption, cell recruitment and, finally, fibrotic encapsulation and extracellular matrix formation. Each of these steps involves a complex cascade of immune modulators including soluble mediators and various cell types. Recent research has focused on the cellular and molecular interactions of the distinct phases of the FBR offering a new basis for therapeutical strategies. The highly dynamic process of the FBR is considerably influenced by the biomaterial composition. Modifications of the type of polymer, the material weight, the filament structure and the pore size are realized and have substantial effects on the in vivo biocompatibility. Moreover, modern mesh technology aims to utilize the available implants as carrier systems for bioactive drugs. Studies in animal models account for the efficiency of these drugs that aim to reduce mesh-related infections or to minimize FBR by influencing inflammation or extracellular matrix remodelling. A thorough understanding of the molecular mechanisms of FBR provides a sophisticated background for the development of new biomaterials at least as carrier systems for bioactive reagents to reduce inflammation and to improve clinical outcome.

Quantitative Characterization of Macrophage, Lymphocyte, and Neutrophil Subtypes Within the Foreign Body Granuloma of Human Mesh Explants by 5-Marker Multiplex Fluorescence Microscopy.

Foreign bodies such as fibers of a surgical mesh induce a typical reaction with an inflammatory infiltrate that forms a surrounding granuloma. This infiltrate is dominated by macrophages, lymphocytes, and neutrophils, whereas its extent of collaboration is widely unknown. In this study, we analyzed 12 samples of surgical meshes explanted from humans by multiplex analyses with three different 5-marker panels - 1. macrophage panel: CD68, CD86, CD105, CD163, and CD206; 2. lymphocyte panel: CD3, CD4, CD8, CD20, and CD68; and 3. neutrophil panel: CD15, histone, MPO, NE, and CD68. Measurement of fluorescence intensity within nuclear masks resulting from DAPI nuclear staining allows exact quantification of cells considered "positive" at a user-defined mean intensity threshold of > 100. Obviously, however, there is no natural threshold as a biological criterion for an intensity that separates "positive" stained cells from unstained cells ("negative"). Multiplex staining of 5 markers always reveals a high rate of coexpression for almost all of the 25 possible marker combinations (= 32 combinations, when using 5 markers simultaneously). The present staining results demonstrate that various morphological and functional subtypes of macrophages, lymphocytes, and neutrophils are abundant in the foreign body granuloma (FBG), which were investigated by regions of interest (ROI) with an area of 1 mm2. The widespread coexpression of two or more markers underscores the complex collaboration network of the inflammatory infiltrate. The ability to combine spatial distribution with exact numerical analysis may offer new perspectives for our understanding of the complex interactions in this multidimensional process.