Characterization of early recurrences following liver resection by ALPPS and two stage hepatectomy in patients with colorectal liver-metastases and small future liver remnants; a translational substudy of the LIGRO-RCT.

BACKGROUND: Associated liver partition and portal vein ligation in staged hepatectomy (ALPPS) is an alternative resection method to portal vein embolization (PVE) in patients with small future liver remnants (FLR) but has been associated with early tumor recurrences. METHODS: Twenty-four patients with colorectal liver metastases (CRLM) patients from the randomized multicenter LIGRO trial comparing outcome of ALPPS (n = 13) vs PVE (n = 11) were included in the study. Mutational analyses of the KRAS, NRAS, BRAF, PIC3CA and TP53 genes of the metastases were performed in 21 patients and correlated to early tumor recurrence. RESULTS: Within 12 months, 13 patients experienced recurrences (6 in TSH group and 7 in ALPPS group). Nine of 13 patients with recurrences had mutations in the TP53 gene, while 3 of 8 patients without recurrence carried the same mutation. Only sporadic cases of the other mutations studied were identified. CONCLUSIONS: ALPPS did not appear to be associated with higher rate of rapid recurrences than PVE following radical resection of colorectal liver metastases. Mutations in genes associated with negative oncologic outcome after surgical resection most likely play a role for tumor recurrences in these patients.

Resection of synchronous liver metastases between radiotherapy and definitive surgery for locally advanced rectal cancer: short-term surgical outcomes, overall survival and recurrence-free survival.

AIM: There is debate as to the correct treatment algorithm sequence for patients with locally advanced rectal cancer with liver metastases. The aim of the study was to assess safety, resectability and survival after a modified 'liver-first' approach. METHOD: This was a retrospective study of patients undergoing preoperative radiotherapy for the primary rectal tumour, followed by liver resection and, finally, resection of the primary tumour. Short-term surgical outcome, overall survival and recurrence-free survival are reported. RESULTS: Between 2009 and 2013, 45 patients underwent liver resection after preoperative radiotherapy. Thirty-four patients (76%) received neoadjuvant chemotherapy, 24 (53%) concomitant chemotherapy during radiotherapy and 17 (43%) adjuvant chemotherapy. The median time interval from the last fraction of radiotherapy to liver resection and rectal surgery was 21 (range 7-116) and 60 (range 31-156) days, respectively. Rectal resection was performed in 42 patients but was not performed in one patient with complete response and two with progressive metastatic disease. After rectal surgery three patients did not proceed to a planned second stage liver (n = 2) or lung (n = 1) resection due to progressive disease. Clavien-Dindo ≥Grade III complications developed in 6.7% after liver resection and 19% after rectal resection. The median overall survival and recurrence-free survival in the patients who completed the treatment sequence (n = 40) were 49.7 and 13.0 months, respectively. Twenty of the 30 patients who developed recurrence underwent further treatment with curative intent. CONCLUSION: The modified liver-first approach is safe and efficient in patients with locally advanced rectal cancer and allows initial control of both the primary tumour and the liver metastases.

Single-centre experience of laparoscopic pancreatic surgery.

BACKGROUND: Laparoscopic resection is regarded as safe and feasible in selected patients with benign pancreatic tumours. Few data exist on laparoscopic surgery for malignant lesions and larger neoplasms in unselected patients. METHODS: The study included all patients admitted to Oslo University Hospital, Rikshospitalet, from March 1997 to March 2009 for surgery of lesions in the body and tail of the pancreas, and selected patients with lesions in the pancreatic head, who underwent surgery by a laparoscopic approach with curative intent. RESULTS: A total of 166 patients had 170 operations, including 138 pancreatic resections, 18 explorations, nine resections of peripancreatic tissue and five other therapeutic procedures. Four patients had repeat procedures. There were 53 endocrine tumours (31.0 per cent), 28 pancreatic carcinomas (16.4 per cent), five cases of metastases (2.9 per cent), 48 cystic tumours (28.1 per cent) and 37 other lesions (21.6 per cent). The total morbidity rate was 16.5 per cent. Fistula was the most common complication (10.0 per cent). Three patients needed reoperation for complications. There were three hospital deaths (1.8 per cent). Median hospital stay following surgery was 4 days. CONCLUSION: Laparoscopic resection of lesions in the body and tail of the pancreas in an unselected patient series was safe and feasible, and should be the method of choice for this patient group in specialized centres.

[Laparoscopic resection of malignant liver tumors: immediate and long-term results].

The study was concerned with laparoscopic liver resection for cancer (140) (1998-2009). Faulty intraoperative performance and postoperative complications were evaluated according to Clavien and Satava. 188 resections were performed in the course of 154 operations. Median operation duration and blood loss were 180 min and 300 ml, respectively. Most patients started taking water on the same day and eating the next day. There were 16 incidents including 6 (3.9%) cases involving change of surgical strategy. Surgical complications were reported in 22 (14.3%) cases including one death from multiple organ failure. Median postoperative intensive care duration and inpatient stay were zero and 3 days, respectively; median follow-up--24-105 months; 3-5 year survival--(68 +/- 6%) and (46 +/- 8%), respectively. 3-year survival in patients with colorectal metastasis to liver, metastasis of neuroendocrine tumors of the gastrointestinal tract and those of hepatocellular carcinoma was 71, 50 and 47%, respectively. Laparoscopic liver resection is a safe and effective method of tumor treatment. It is least invasive and followed by good end results.

[Laparoscopic resection of primary and metastatic malignant tumors of the adrenals].

An analysis was made of experience with treatment of 24 patients who underwent laparoscopic adrenalectomy for adrenocortical carcinomas (in 7 patients) and metastases in adrenals (in 17 cases). Laparoscopic adrenalectomy was shown to be a safe and effective method of treatment of primary and metastatic tumors of the adrenals. The method can replace open operative intervention in the majority of patients with metastases to adrenals and primary cancer of the adrenals.

Associating liver partition and portal vein ligation for staged hepatectomy in patients with colorectal liver metastases--Intermediate oncological results.

BACKGROUND: Colorectal liver metastases (CRLM) not amenable for resection have grave prognosis. One limiting factor for surgery is a small future liver remnant (FLR). Early data suggests that associating liver partition and portal vein ligation for staged hepatectomy (ALPPS) effectively increases the volume of the FLR allowing for resection in a larger fraction of patients than conventional two-stage hepatectomy (TSH) with portal vein occlusion (PVO). Oncological results of the treatment are lacking. The aim of this study was to assess the intermediate oncological outcomes after ALPPS in patients with CRLM. MATERIAL AND METHODS: Retrospective analysis of all patients with CRLM operated with ALPPS at the participating centres between December 2012 and May 2014. RESULTS: Twenty-three patients (16 male, 7 female), age 67 years (28-80) were operated for 6.5 (1-38) metastases of which the largest was 40 mm (14-130). Six (27.3%) patients had extra-hepatic metastases, 16 (72.7%) synchronous presentation. All patients received chemotherapy, 6 cycles (3-25) preoperatively and 16 (70%) postoperatively. Ten patients (43%) were rescue ALPPS after failed PVO. Severe complications occurred in 13.6% and one (4.5%) patient died within 90 days of surgery. After a median follow-up of 22.5 months from surgery and 33.5 months from diagnosis of liver metastases estimated 2 year overall survival was 59% (from surgery) and 73% (from diagnosis). Liver only recurrences (n = 8), were treated with reresection/ablation (n = 7) while lung recurrences were treated with chemotherapy. CONCLUSION: The overall survival, rate of severe complications and perioperative mortality associated with ALPPS for patients with CRLM is comparable to TSH.

Reduced CD4 cell counts in blood do not reflect CD4 cell depletion in tonsillar tissue in asymptomatic HIV-1 infection.

OBJECTIVE: To investigate whether the loss of CD4 cells seen in peripheral circulation of HIV-1-positive individuals reflects a similar depletion of CD4 cells from lymphoid tissue. DESIGN: CD4 and CD8 cells in tonsillar mononuclear cell suspensions were quantified relative to tonsillar B cells, as these were thought to remain numerically unchanged in the course of HIV infection. Results were related to the CD4 cell counts in blood and to the clinical status of the patients. METHODS: Blood samples and tonsillar tissue were obtained from 13 HIV-1-seropositive individuals and six seronegative controls. B cells and T-cell subsets in mononuclear cells were quantified using a three-colour flow cytometry protocol. Histological sections were morphologically classified and B-cell areas were quantified by morphometry. RESULTS: The B-cell fraction was confirmed to be relatively unchanged in asymptomatic HIV-1-seropositive individuals compared with controls. The tonsillar CD4 : B-cell ratios in asymptomatic individuals was similar to those seen in controls, whereas the CD4 : B-cell ratios in symptomatic HIV-1-infected individuals were greatly reduced. The tonsillar CD4 : CD8 cell ratios in HIV-1-infected individuals were much lower than those seen in controls, in the asymptomatic group due to a considerable expansion of the tonsillar CD8 cell subset, and in the symptomatic group also due to a loss of CD4 cells. CONCLUSIONS: We found no evidence of CD4 cell depletion in tonsillar tissue in asymptomatic HIV-1-infected individuals despite low CD4 cell counts in blood. Loss of CD4 cells from this lymphoid tissue seems to occur as a late-stage phenomenon correlated with the onset of clinical symptoms.

Deregulation of the CD95/CD95L system in lymphocytes from patients with primary acute HIV infection.

OBJECTIVE: To analyze the role of CD95/CD95 ligand (CD95L) expression and functionality in peripheral blood lymphocytes (PBL) during primary, acute HIV syndrome (AHS) and in the subsequent period. PATIENTS: Twelve patients were studied during the acute phase of the viral infection and most were followed for some months. METHODS: Cell culture and cytotoxicity assays based upon 51Cr release and flow cytometry were used to evaluate cell killing via CD95 molecule, flow cytometry to assess surface antigens, enzyme-linked immunosorbent assay (ELISA) for the determination of soluble CD95 and CD95L plasma levels, quantitative competitive (QC) reverse transcription polymerase chain reaction (RT-PCR) with an original RNA competitor for the analysis of CD95L mRNA expression and QC RT-PCR for determining plasma viral load. RESULTS: The analysis of PBL during this phase revealed that almost all cells, including CD8 T cells with a virgin phenotype, B lymphocytes and natural killer cells displayed CD95 molecules on the plasma membrane. Activation of CD95 on the surface of isolated lymphocytes by anti-CD95 monoclonal antibodies or binding to CD95L induced rapid apoptosis. However, CD95L mRNA was not expressed in PBL from these patients and was poorly inducible. Soluble CD95 was found in the plasma of all patients, but only in a few at high levels, even some months after seroconversion. In contrast, soluble CD95L was detected in only one patient, this occurring after the symptomatic period. For 10 of the 12 patients, expression of CD95 on the cell membrane or in the plasma did not correlate with the plasma viral load, which varied widely from patient to patient. Further, plasma levels of soluble CD95 were not altered by decreased lymphocyte activation or by efficient antiretroviral therapy. CONCLUSIONS: In patients experiencing an acute, primary HIV infection, a prolonged deregulation of the CD95/CD95L system may exist, which is probably not entirely related to virus production but may contribute to the pathogenesis of the disease. The hypothesis can be put forward that a complex balance exists between proapoptotic events (increase in CD95 expression), probably triggered by the host as a method to limit viral production, and antiapoptotic events (decrease in CD95L expression) probably triggered by the virus as a way to increase its production and survival.

Characterization of HIV type 1 from Romanian children: lack of correlation between V3 loop amino acid sequence and syncytium formation in MT-2 cells.

The biological properties and amino acid sequences of the third variable domain (V3 loop and flanking regions) of the env region of 34 HIV-1 isolates obtained from Romanian children were analyzed. Unambiguous nucleic acid sequences were obtained from 31 isolates. The derived V3 amino acid sequences were highly homologous (93-100%) and clustered with the HIV-1 subtype F Romanian consensus. Five of the 31 isolates presented a syncytium-inducing phenotype in MT-2 cells and established continuous viral replication in various CD4+ cell lines (rapid/high phenotype). The V3 sequence from one of these isolates showed a slightly lesser degree of homology with the consensus sequence. The presence of positively charged amino acids at positions 306 and 320 has been strongly associated with the ability to induce syncytia in MT-2 cells, whereas negatively or uncharged amino acids at these positions are present in non-syncytium-inducing isolates (slow/low phenotype). There was, however, no correlation between phenotype and amino acid sequence in the five syncytium-inducing isolates; negatively or uncharged amino acids were conserved at positions 306 and 320 for all 31 isolates in sequences obtained from PBMCs. A tendency toward a more positive net charge in the V3 loop of syncytium-inducing isolates was noted. These data confirm the recent observations that HIV-1 isolates from Romania not only cluster in subtype F, but also show a high degree of interpatient homogeneity in the V3 region.(ABSTRACT TRUNCATED AT 250 WORDS)

CD8+ T cells from HIV type 1-seronegative individuals suppress virus replication in acutely infected cells.

CD8+ lymphocytes (CD8 cells) have been shown to inhibit replication of the human immunodeficiency virus (HIV) in vitro when cocultured with HIV-infected CD4+ lymphocytes (CD4 cells). This suppressive effect on HIV replication in experimentally infected CD4 cells has so far been demonstrated only for CD8 cells from HIV-seropositive individuals. In the present study we have investigated if CD8 cells from HIV-negative individuals can also suppress HIV replication in experimentally infected CD4 cells. Positively selected CD4 cells were infected with phenotypically different primary isolates of HIV type 1 and 2 (HIV-1 and HIV-2). Graded numbers of CD8 cells were added to the infected cultures. The T cells were activated by antibodies directed against the CD3 molecule or the T cell receptor. Culture supernatants were harvested for HIV p24 quantitation and the CD8 suppression of HIV replication was calculated by comparing p24 levels from parallel cultures in the presence or absence of CD8 cells from different donors. We show that CD8 cells from unexposed HIV-seronegative blood donors are able to control HIV-1 and HIV-2 replication in experimentally infected autologous CD4 cells. The antiviral activity of CD8 cells from and HIV-naive individual was reproducible over time and the suppressive effect was comparable to that seen with CD8 cells from HIV-positive individuals. The infected cells were not eliminated from the cultures. The suppressive effect of CD8 cells varied depending on the dose and biological phenotype of the virus used for infection. Thus, exposure to HIV in vivo is not a prerequisite for CD8 cells to exert a suppressive effect on HIV replication in acutely infected cells.

Normalization of CD4+ cell numbers and reduced levels of memory CD8+ cells in blood and tonsillar tissue after highly active antiretroviral therapy in early HIV type-1 infection.

Antiretroviral therapy increases the number of both CD4+ and CD8+ T cells in the blood of HIV-1-positive patients with advanced disease. In the present study, we have examined the kinetics of CD4+ and CD8+ T cell restoration in blood and lymphoid tissue in asymptomatic HIV-1-positive individuals with high CD4+ cell counts during highly active antiretroviral treatment. Tonsillar biopsies and blood samples were collected at baseline and at regular intervals during the following 48 weeks and from HIV-1-negative controls. Mononuclear cells from blood and tonsils were phenotyped and quantified by three-color flow cytometry. After 48 weeks of therapy, blood CD4+ cell counts in the HIV-1-infected group were comparable to those found in uninfected controls. Naive CD4+ T cells in blood increased during the initial 2 weeks in parallel with reduced plasma viremia. Both naive and memory CD4+ T cells in blood reached normal numbers by week 48, whereas the CD4+ naive/memory cell ratio in tonsils was within normal range throughout the study. The level of memory CD8+ T cells in blood declined during the first 8 weeks in parallel with a reduction in the tonsillar memory CD8+ T cells. Naive CD8+ T cells in the blood increased after 4 weeks, while the level of naive CD8+ T cells in tonsils remained unaltered. Our data indicate that in the early stages of HIV-1 infection antiretroviral therapy normalizes CD4+ cell counts and causes a decrease in the level of memory CD8+ cells in blood and lymphoid tissue, suggesting reduced CD8+ cell turnover in response to reduced viral replication.

Correlates of apoptosis of CD4+ and CD8+ T cells in tonsillar tissue in HIV type 1 infection.

The immunopathogenesis of human immunodeficiency virus type 1 (HIV-1) infection has been associated with increased death by apoptosis of T cell subsets. In the present study, we have examined correlates of apoptosis of CD4+, CD8S+CD28+, and CD8+CD28- T cells in tonsillar lymphoid tissue in persons with HIV-1. Single-cell suspensions of tonsillar lymphocytes were analyzed by flow cytometry to determine the fraction of cells showing typical characteristics of apoptosis as well as the expression of activation markers within the live and the apoptotic cell populations. The proportion of cells carrying infectious provirus was quantified by limiting dilution analysis. Compared with uninfected controls, apoptosis of both CD4+ and CD8+ T cells was enhanced in HIV-1 infection and was higher among CD8+ than among CD4+ T cells. Apoptosis of CD28-cells was more prevalent than apoptosis of CD28+ cells for both CD4+ and CD8+ T cells. Occurrence of apoptosis of CD4+ T cells correlated with provirus levels and proportional expression of the activation marker HLA-DR. Apoptosis of CD8+CD28+ cells correlated with expression of the activation markers CD69 and HLA-DR while apoptosis within CD8+CD28- cells did not correlate with any of the studied parameters. Although apoptosis was much more prevalent among CD8+ than CD4+ T cells, CD8+ T cells still accumulated in tonsillar lymphoid tissue in persons with HIV-1. Our data may be interpreted to suggest that apoptosis of CD4+, CD8+CD28+, and CD8+CD28- cells in tonsillar tissue is regulated by different mechanisms and the results are of importance to our understanding of the immunopathogenesis of HIV-1 infection.

Changes in tonsillar tissue in early HIV-1 infection and during 3 years of antiretroviral therapy.

Tonsillar tissue from individuals in the early stages of HIV-1 infection was studied during the natural course of infection and during antiretroviral therapy with and without a protease inhibitor in order to investigate markers of clinical progression and evaluate the effects of therapy. Tonsillar biopsies and blood samples were collected at regular intervals during 3 years and clinical observations were noted. Tonsillar morphology was evaluated and the fragmentation of the follicular dendritic cell network was quantified by standardised follicular fragmentation rate (FR) analysis. Lymphocyte subsets were phenotyped by flow cytometry, and viral load was calculated by limiting dilution assay. The FRs were higher in the HIV-1-infected individuals than in the uninfected controls, although tonsillar tissue from both groups contained follicular fragmentation. During HIV-1 infection, the FR increased and the tonsillar CD4/CD8 ratio declined. During maximum viral suppression, FR approached that of controls while tonsillar T cell subsets and blood CD4 cell counts normalised. Even when virus suppression was incomplete, tonsillar improvements were observed in parallel with a resolution of the HIV-1-related dermatological disorders. However, persistent viral replication paralleled distortion of the tonsillar architecture. We suggest that a normalisation of the lymphoid tissue may have important functional and clinical implications in HIV-1 infection.

Release of bicarbonate from damaged and restituted gastric mucosa in the cat.

BACKGROUND: Gastric mucosal damage leads to luminal alkalinization, but its dependence on mucosal blood flow and acid secretory state of the mucosa is not known. This study examined release of bicarbonate to the gastric lumen and mucosal blood flow in cats after mucosal damage caused by 2 mol/L NaCl and during 90 minutes of epithelial restitution. METHODS: Bicarbonate was calculated from measurements of pH and PCO2 in the luminal perfusate. Mucosal blood flow was measured with microspheres. RESULTS: Luminal bicarbonate increased more than twofold after damage in pharmacologically nontreated, pentagastrin-treated, and omeprazole-treated animals (P < 0.001). Luminal bicarbonate thereafter decreased completely to pre-damage level in pentagastrin-treated, partly in nontreated, but remained elevated in omeprazole-treated animals. Mucosal blood flow increased about 100% 15 minutes after damage (P < 0.001), irrespective of secretory state. Bicarbonate availability (arterial [HCO(3-)] x mucosal blood flow) was significantly related to luminal release of bicarbonate from the newly damaged (P < 0.01) but not from the restituted mucosa. CONCLUSIONS: (1) From the newly damaged mucosa, the luminal release of bicarbonate is related to availability of blood-borne bicarbonate. (2) From acid-stimulated restituted mucosa, the bicarbonate produced by the parietal cells is not released to the lumen, but either consumed within the mucosa by back-diffusing H+ or distributed to the systemic circulation.

Activation and proliferation of CD8+ T cells in lymphoid tissues of HIV-1-infected individuals in the absence of the high-affinity IL-2 receptor.

Activation of CD8+ T cells may have important pathogenic implications in HIV-1 infection. Studies of this process have so far been confined to cells from the peripheral blood. In the present study, we have examined molecules involved in activation and proliferation of CD8+ T cells in lymphoid tissues from HIV-1-infected patients. Tonsillar tissue and blood samples from 13 HIV-1-infected patients and 6 seronegative controls were examined for cell surface expression and the presence of mRNA for CD69, CD25, and HLA-DR. Intonsillar tissue, the number of CD8+ T cells was increased several fold in HIV-1-infected patients compared with controls. The majority of these cells expressed CD69 and HLA-DR, but virtually no tonsillar CD8+ T cells were found to express CD25 on the cell surface or at the mRNA level. Following in vitro activation, however, almost all activated CD8+ T cells were found to express CD25. Tonsillar CD4+ T cell numbers were maintained or reduced compared with controls, and a considerable proportion expressed CD25. The data suggest that CD8+, but not CD4+ T cells proliferate extensively in lymphoid tissues in HIV-1-infected patients in the absence of the high-affinity interleukin-2 (IL-2) receptor.

Dynamics of HIV-1 replication following influenza vaccination of HIV+ individuals.

Levels of HIV-1 have been reported to increase in peripheral blood after influenza vaccination of HIV+ individuals. In this study we have evaluated the dynamics of these changes. Ten HIV-1+ individuals classified in revised CDC clinical categories B and C as well as five seronegative healthy controls were vaccinated with the recommended influenza strains. HIV viral RNA and proviral DNA were sequentially quantified in serum and blood lymphocytes, respectively. Nine of the 10 HIV+ individuals had an increase in the frequency of infected CD4 cells 2 weeks after influenza vaccination. Individuals with low viral load had a rapid increase in viraemia and a small increase in frequency of infected cells in peripheral blood. In contrast, individuals with high viral load had a small drop in viraemia followed by a significant rise in the rate of infected cells. The observed change may resemble those taking place during intercurrent infections in HIV+ individuals. The effects of the relative increases in infectious virus after the transient viraemic phase should be further investigated to evaluate potential risks of vaccination.

Gastric bicarbonate secretion, acid secretion, and mucosal blood flow during influence of pentagastrin and omeprazole in the cat.

In this study secretion of bicarbonate and acid and mucosal blood flow were determined simultaneously in cats. The gastric lumen of anesthetized cats was continuously perfused with isotonic saline. Secretion of HCO-3 and H+ was calculated from continuous measurements of pH and PCO2 in the perfusate. Mucosal blood was measured by means of radiolabeled microspheres. Under resting acid secretory conditions, bicarbonate secretion into the gastric lumen averaged 1.0 mumol/min. Somewhat surprising, both omeprazole (4 mg/kg as bolus) and pentagastrin (16 micrograms/kg.h intravenously) significantly reduced the HCO-3 secretion. Omeprazole did not influence mucosal blood flow, whereas corpus mucosal blood flow increased during pentagastrin stimulation. Under resting acid secretory conditions and during omeprazole treatment there was a close linear relationship between acid and bicarbonate secretion. No such relationship was found during pentagastrin stimulation of the mucosa. No consistent relationship was obtained between blood flow and bicarbonate secretion in normal gastric mucosa.

Correlates of latent and productive HIV type-1 infection in tonsillar CD4(+) T cells.

Correlates of virus load and characteristics of virus-producing cells in tonsillar tissue were investigated. Our results suggest that when less than 1:100 tonsillar CD4(+) T cells from individuals infected with HIV type-1 (HIV-1) contain replication competent provirus, the level of CD4(+) T cells in tonsils is comparable to that observed in uninfected individuals. Virus load at or above this level was associated with low CD4 cell numbers in tonsillar tissue. Only a few percent of all infected T cells in tonsillar tissue were active virus producers, with minor differences observed between individuals. Plasma viremia was found to correlate with infectious virus load in tonsillar tissue. With less than 1:1,000 of CD4 cells in lymphoid tissues being involved in active virus production, direct cytopathic effect by HIV-1 on infected CD4 cells is unlikely to fully explain the immunodeficiency seen in AIDS.