Radioembolisation with (90)Y-labelled resin microspheres in the treatment of liver metastasis from breast cancer.

OBJECTIVES: Metastatic breast cancer is a heterogeneous disease, commonly affecting the liver. We report our experience with (90)Y radioembolisation (RE) and its effects on the survival of patients with treatment-refractory breast cancer liver metastases. METHODS: A total of 77 female patients affected by breast cancer were accepted into our department for RE. Inclusion criteria were inoperable and chemotherapy-refractory hepatic metastases, acceptable performance status, sufficient residual liver, no significant hepato-pulmonary shunts. Patients were divided in two groups: group 1 (29 patients) included those with Eastern Cooperative Oncology Group (ECOG) score 0, liver involvement (0-25 %) and no extrahepatic disease (EHD); group 2 (23 patient) included patients with ECOG score 1-2, liver involvement (26-50 %) and evidence of EHD. RESULTS: A total of 25 patients were considered ineligible. The median age of the remaining 52 patients was 57.5 years. The median overall survival was 11.5 months and better in those whose performance status and liver function were preserved (14.3 versus 8.2 months). According to Response Evaluation Criteria in Solid Tumor (RECIST), partial response (PR) was achieved in 29 patients (56 %), stable disease (SD) was achieved in a further 18 patients (35 %) and 5 patients showed progressive disease (PD) (10 %). DISCUSSION: (90)Y RE is effective in the treatment of liver metastases from breast cancer. We demonstrated a relevant survival and encouragingly high response rate in patients with treatment-refractory disease.

Radioembolisation using yttrium 90 (Y-90) in patients affected by unresectable hepatic metastases.

PURPOSE: This study was done to evaluate the effectiveness of radioembolisation of liver metastases with yttrium 90 (Y-90) in patients with no response to chemotherapy. MATERIALS AND METHODS: From February 2005 to January 2008, we treated 110 patients affected by liver metastatic disease from colorectal, breast, gastric, pancreatic, pulmonary, oesophageal and pharyngeal cancers and from cholangiocarcinoma and melanoma. We excluded patients with bilirubin level >1.8 mg/dl and pulmonary shunt >20% but not patients with minor extrahepatic metastases. RESULTS: We obtained a complete /partial response in 45 patients, stable disease in 42 patients and progressive disease in 23 patients. In 90 cases, we obtained a decrease in specific tumour marker level. The technical success rate was 96%, and technical effectiveness estimated at 3 months after treatment was 83.6%. Side effects were grade 4 hepatic failure in one case, grade 2 gastritis in six cases and grade 2 cholecystitis in two cases. The median survival and progression-free survival calculated by Kaplan-Meier analysis were 323 days and 245 days, respectively. CONCLUSIONS: According to our 3-year experience, Y-90 radioembolisation (SIR-spheres) is a feasible and safe method to treat liver metastases with an acceptable level of complications and a good response rate.

[Massive haemoperitoneum due to traumatic rupture of multifocal hepatocarcinoma in the right hepatic lobe. Case report]. / Emoperitoneo massivo da rottura di epatocarcinoma multifocale del lobo destro. Case report.

UNLABELLED: Abdominal blunt trauma is the main cause of death in people younger than 40 years old. The liver injury still represents a challenging problem. Isolated hepatic injury is rare and it occurs more frequentely in polytraumatizated patients and causes massive haemoperitoneum. The Authors report a case of a 83 years-old woman admitted to Emergency Department for syncope due to an active bleeding arising from a rupture of a right hepatic lobe unsuspected tumor. The computer tomography (CT) scans showed a clear pattern of liver laceration of the VI segment with contrast enhancement spreading in the surrounding tissues, and detected a multifocal hepatocarcinoma located in the VI, VII and VIII segments. Patient's haemodinamically unstable conditions suggested an urgent laparotomy. An accurate perihepatic packing with sterile-drape were successfully employed to control liver hemorrage. Temporary abdominal closure, followed by hepatic arteriography and the right hepatic artery embolization, completed the damage control. Re-exploration laparotomy after 72 hours confirmed the definitive haemostasis and the pack removal was performed without complications. CONCLUSIONS: CT plays a leading role in the diagnosis of liver damage. The patient's haemodynamic status is the principal criterion determining conservative or operative therapy in blunt liver injury. The early perihepatic packing followed by artheriographic embolization to stop liver hemorrhage showed efficacy and safety for the patient. The packing performed with sterile-drape is able to avoid removal complications and 72 hours timing for the pack removal is effective to avoid re-bleeding.

[A case of arterial chemoembolization for hepatocellular carcinoma with 11 years of survival]. / Un caso di epatocarcinoma trattato con chemio-embolizzazione arteriosa transcatetere con sopravvivenza di 11 anni.

Hepatocellular carcinoma (HCC) is the most common malignancy of the liver and the third most common cause of cancer mortality worldwide. The major risk of developing HCC is associated with HBV and HCV hepatitis. Liver transplant (LT) is the gold standard for "small" HCC (HCCs) in Child-Pugh class A cirrhotic patients. However its use has been restricted by the severe shortage of donors, so that hepatic resection (HR) is often performed in these patients. In the last two decades image-guided interventional catheterization and ablative regional treatment procedures have revolutionized the therapy of unresectable primary and secondary liver tumors. The Authors present a case of a 61-years old man with Child-Pugh class A HCCs. The age and the previous history of bladder carcinoma made the patient not suitable for LT. The patient refused HR so that transarterial chemoembolization combined to thermo-ablation therapy and oral intake of tamoxifen were proposed. Patient's tolerance to the treatments has been good. During 11-year follow-up there was earlier intrahepatic progression of the tumor followed by reduction in size and number of the lesions. In spite of the scarce prognosis, chemoembolization and immunotherapy allowed to achieve a satisfactory local control of disease in our patient and guaranteed a good quality of life at long-term follow-up.

[Massive lower gastrointestinal bleeding due to diverticular disease during antiplatelet therapy. Case report]. / Enterorragia massiva da diverticolosi sigmoidea in corso di terapia antiaggregante. Caso clinico.

Diverticular disease is very frequent in Western countries; in 5% of the cases it is the cause of serious bleeding, haemodynamic instability and death. The authors report a case of 74 years old patient with severe lower gastrointestinal bleeding. She was in antiplatelet treatment with acetylsalicylic acid (100 mg/die) and clopidogrel (75 mg/die) for preventing the restenosis of medicated stents positioned to treat an acute coronary syndrome. At the same time the patient was under treatment for primary hypercholesterolemia with rosuvastatin (20 mg/die). The severe haemorrhage demanded haemodynamic stabilization, achieved by colloid infusion and blood transfusions. The bleeding continued; selective arteriography showed it's origin from the areas of the sigmoid and superior hemorrhoidal arteries. During the procedure, embolization of the inferior mesenteric artery using spiral type BALT was performed, with consequent bleeding interruption. Fifteen days after the embolization, a rectosigmoid colonoscopy showed a sigmoid diverticular disease. The treatment with acetylsalicylic acid and clopidogrel has surely contributed to the severity of the hemorrhage. Recent experimental and clinical evidence suggests a possible antiplatelet effect of the statins.

Incorporation of cultured embryonic cells into transgenic and chimeric, porcine fetuses.

The derivation of murine, embryonic stem cells, and their use in the generation of transgenic mice, are well-established procedures. Application of these methodologies to non-murine species, however, remains to be fully realized. Non-murine embryonic stem cells would be of considerable value to studies in comparative development, and would raise important implications for agriculture and biotechnology. Here we report the achievement of chimerism and transgenesis in the domestic pig (Sus scrofa) at a fetal stage by the embryonic stem-cell methodology, using an embryonic cell line which previously we described as having the capacity to differentiate in vitro. This entailed: transformation, by electroporation, of a cell line, PE1, with the bacterial gene for neomycin-resistance; re-introduction of a polyclonal population of neo-PE1 cells into host blastocysts, followed by transfer of reconstructed embryos into a pseudopregnant recipient; and subsequent derivation of transgenic and chimeric porcine fetuses, as determined by two independent molecular assays of fetal genomic DNA. In the first instance, chimerism was revealed in one fetus by the presence of the transgene, as detected by Southern blotting; and in the second instance, in that and another fetus by the presence of supernumerary alleles for the class II major histocompatibility locus, SLA-DQB*C, by single-strand conformation polymorphic analysis. The contribution of neo-PE1 cells to the first chimeric and transgenic fetus was approximately 25%, and to the second chimeric fetus, below the level of detection by Southern blotting (i.e. less than 10%). The results indicate that, at the time of embryo reconstitution, a proportion of neo-PE1 cells were pluripotent and of the primary ectodermal lineage.

Early changes in embryonic nuclei fused to chemically enucleated mouse oocytes.

Mouse oocytes were chemically enucleated by subjecting them to etoposide and cycloheximide treatment during the first meiotic division (Fulka, Jr. and Moor, Mol. Reprod. Dev. 34:427-430, 1993) and thereafter electrofused to karyoplasts prepared from: (i) two-cell stage embryos at the G2-phase; (ii) four-cell stage blastomeres (S- or G2-phase); or (iii) embryonic stem (ES) cells. In the first series of experiments we used fusion conditions which do not induce egg activation to define the series of nuclear changes that are initiated immediately following fusion. Although fusion is evident within 5-10 min of induction, nuclei remain visible for up to 20 min prior to chromatin condensation and the formation of metaphase plates (60-90 min post fusion). After activation, the anaphase-telophase transition is completed within 1-2 h, followed thereafter by cleavage of 75% of reconstituted eggs into two equal nucleated blastomeres, irrespective of the origin of the nuclei used for fusion. We conclude from the first study that a protocol involving fusion without activation, followed 90 min later by activation, is likely to be optimal for nuclear transplantation using MII-phase cytoplasts. In the second series of experiments the above optimized protocol was used to study the effects of different cell cycle combinations on chromosome organization in eggs reconstituted by nuclear transplantation. Both G1- and S-phase karyoplasts fused to MII-phase cytoplasts exhibited spindle abnormalities in all eggs studied. Characteristic abnormalities in these cell cycle combinations included chromatin fragmentation and joining or aggregations of chromatin.(ABSTRACT TRUNCATED AT 250 WORDS)

Nuclear remodelling and early development in cryopreserved, porcine primordial germ cells following nuclear transfer into in vitro-matured oocytes.

Nuclear transfer was conducted in the pig using, as karyoplasts, primordial germ cells which had been cryopreserved. The cytoplasts were presumptive S- or MII-phase, in vitro-matured oocytes, which had been enucleated mechanically. Enucleation was effective in 94.3% of cases. Karyoplasts were introduced into the perivitelline space, in close contact with the cytoplasts, and the complexes fused by electrical stimulation and activation. Activation was successful in 82-88% of nonmanipulated, pulsed oocytes, and in 55% of germ cell-oocyte complexes. The reconstituted embryos were examined for nuclear remodelling and cleavage in vitro. Nuclear swelling was more prominent when MII-phase cytoplasts, rather than S-phase, cytoplasts, were used. After 24 h in culture, the cleavage rate was not significantly different whether blastomeres or primordial germ cells were used as karyoplasts, and whether MII-phase or S-phase cytoplasts were used. However, after 72 h in culture, the developmental rate was higher when MII-phase cytoplasts (75%) were used for the recipients of blastomeres compared with S-phase cytoplasts (38.5%, p < 0.05). Similar tendencies were observed with germ-cell nuclear transfer when inositol was used as medium for electrofusion (60% vs 27.8%, p < 0.05). Furthermore, when MII-phase cytoplasts were used, the nuclear transferred embryos derived from blastomeres developed at a significantly higher rate than from primordial germ cells (37.5%, p < 0.05). We conclude that cryopreserved primordial germ cells are competent to undergo nuclear remodelling and cleavage during 72 h or incubation in vitro to the 4-cell stage, following nuclear transfer to enucleated, activated (S-) or MII-phase oocytes. This experimental system may help to elucidate events in the early development of pig embryo following nuclear transfer using germ cells as karyoplasts.

Retinoids are endogenous to the porcine blastocyst and secreted by trophectoderm cells at functionally-active levels.

Retinoic acid and its isomers are the major morphogens in vertebrate development. For mammals, it was previously considered that circulating retinoic acid was recruited from the uterine environment, to influence embryonic differentiation, morphogenesis and development. Here we report that retinoic acid is endogenous to the blastocyst of the domestic pig, Sus scrofa, as detected by high pressure liquid chromatography. Furthermore, using a continuous, normal line of porcine trophectoderm cells, TE1, we have identified the trophoblast as a major source of retinoids. Endogenous retinoic acid was found at a concentration of approximately 35 nM in extracts of entire blastocysts, and at a similar concentration in extracts of TE1 cells. Retinoids in explants of blastocysts and in conditioned medium from the TE1 cell line were found to be functionally-active, inducing gene expression from a retinoic-acid-responsive enhancer element in an in vitro assay system. In conclusion, we propose that there is a morphogenetic role for endogenous, and trophoblast-derived, retinoids in the early development of the pre-gastrulation porcine embryo; and that the TE1 cell line therefore provides a useful in vitro system for the study of retinoid metabolism. Furthermore, an implication of this study is that endogenous retinoids may play an active role in the pre-implantation embryology of other species, such as the human.

Parietal endoderm cell line from a rat blastocyst.

We have derived a cell line, RE1, from a pre-implantation rat blastocyst, resembling morphologically the L2 cell line from a parietal yolk sac carcinoma of the rat, as well as parietal endoderm cell lines of the mouse. The sub-cellular organization and epithelial characteristics of RE1 cells are described. The cells express cytokeratins of simple epithelia, and vimentin; and demonstrate synthesis of proteins of the extracellular matrix, such as laminin and collagen IV. Extensive Reichert's-like basement membrane is formed by RE1 cells when grown in suspension as aggregates. Cells have a microvillous surface morphology and abundant, rough endoplasmic reticulum which is swollen with apparent secretory material. These morphological and cytochemical features are characteristic of parietal endoderm cells in vivo, and the RE1 cell line is deduced to be rat parietal endoderm. In addition, RE1 cells were examined for expression of stage-specific embryonic antigens: cells reacted with antibody against SSEA-1/TEC-1 and EMA-1, constituting the first observation of parietal endoderm cells expressing the respective epitopes. RE1-cell monolayers did not generate transepithelial resistances or potential differences in vitro, consistent with their formation of leaky epithelia. Our observations on RE1-cell morphology and ultrastructure are consistent with the occurrence of epithelial-mesenchyme transitions in culture.

Electrogenic sodium transport mediated by an amiloride-sensitive conductance in a porcine trophectoderm cell line.

This study examined the electrical properties and Na+ transport function of a porcine trophectoderm cell line, TE1, which forms a polarized epithelium in culture. Specifically, the capacity of TE1 cells to generate a transepithelial potential difference, and to modify selectively the Na+, K+ and Cl- ionic composition of medium in the apical and basolateral compartments, was examined over a 48-h period using monolayers cultured on permeable tissue culture supports. TE1-cell monolayers formed 'tight epithelia' in that significant transepithelial electrical resistances (RT; median value 5.30 k(ohms)/cm2, range 2.26-9.18 k(ohms)/cm2, n = 72), and electrical potential differences (VT; maximum mean value at 24 h, 42.9 mV, SEM 7.14; n = 6) were generated. It is concluded that: (1) the VT was generated by the amiloride-sensitive, Na+ absorptive function; (2) K+ transport across the monolayers was related to the electrogenic Na+ transport function; and (3) the three ions traverse the epithelium by active transport or co-transport, rather than simple diffusion. These data substantiate the proposed role for porcine trophectoderm of generating electrical and chemical potentials in vivo, and for regulating the environment of the blastocoel cavity in the pre-implantation porcine embryo.

Isolation and characterization of a feeder-dependent, porcine trophectoderm cell line obtained from a 9-day blastocyst.

We have established in culture a feeder-dependent cell line, termed TE1, from a 9 day, pre-implantation, porcine embryo. TE1 cells were observed by light and electron microscopy, and characterized by immunocytochemistry: the morphology, cytology and ultrastructure of this cell line are described. The cells display epithelial characteristics, as revealed using immunofluorescence microscopy with antibody against cytokeratins of simple epithelia, but not with antibody against vimentin. The cells demonstrate many morphological and cytochemical features in common with trophectoderm of the intact porcine blastocyst. For example, TE1 cells are polarized and possess tight junctions at their borders, similar to those found in trophectoderm of the pre-implantation embryo. Moreover, TE1 cells label positively for the porcine trophectoderm-specific monoclonal antibody, SN1/38. Thus, by several important criteria TE1 is deduced to be a porcine trophectoderm cell line.

Radiation therapy of spinal metastases: results with different fractionations.

AIMS AND BACKGROUND: Local radiotherapy plays an important role in the palliative treatment of all skeletal metastases, particularly those of the spine, with the purpose to obtain pain relief and prevent pathologic fractures or vertebral collapse. METHODS: From June 1991 to October 1993, 95 patients with a total of 103 sites of spinal metastases were treated at the Institute of Radiology of the University of Rome "La Sapienza". Fractionations and total doses were divided as single fractions of 800 cGy, hypofractionated multiple fractions for a total dose of 20 Gy administered in 4-5 days, and conventional multiple fractions for a total dose of 30-40 Gy in 2-4 weeks. An evaluation of the efficacy of the different radiation treatments was performed with the use of a simplified descriptive pain scale. RESULTS: Seventy-three (70.9%) of 103 treatments were evaluables. An overall response rate of 82.2% was obtained: complete in 38.3% and partial in 43.8%, irrespective of total dose, fractionation and location of irradiated spinal metameres. The analysis of results did not show significant differences between the treatment courses. CONCLUSIONS: We confirm that radiation therapy has a major role in the management of pain control and prevention of fractures in patients with spinal metastases. Hypofractionated and single fraction treatments showed equal efficacy compared to more prolonged therapy, with an advantage for the patient and the radiation therapy institution.

Poly(ADP-ribosyl)ation of a herpes simplex virus immediate early polypeptide.

This paper reports in vitro poly(ADP-ribosyl)ation of the herpes simplex virus type 1 (HSV-1) immediate early polypeptide Vmw175. The phenomenon was most clearly observed by use of the temperature-sensitive mutant tsK, which overproduces Vmw175 at the nonpermissive temperature (NPT) and has a mutation in the coding sequences for this polypeptide. Nuclei prepared from cells which were infected with tsK at NPT and subsequently downshifted to the permissive temperature incorporated [32P]NAD into Vmw175. This reaction did not occur when nuclei were prepared from cells constantly maintained at NPT, showing that only functional Vmw175 can be radiolabeled with [32P]NAD. The identity of the acceptor protein was confirmed by demonstrating the expected electrophoretic mobility differences between the HSV-1 and HSV-2 counterparts of Vmw175. The use of suitable inhibitors demonstrated that the reaction represented mono- or poly(ADP-ribosyl)ation, and further analysis showed the presence of long poly(ADP-ribose) chains attached to Vmw175. Poly(ADP-ribosyl)ation may be important as a cause or result of the regulation of viral transcription by Vmw175. Radiolabeling of another virus-specified polypeptide (approximate molecular weight 38,000), thought to be a structural component of the input virus, is also reported.

Premeiotic disruption of duplicated and triplicated copies of the Neurospora crassa am (glutamate dehydrogenase) gene.

Premeiotic inactivation of duplicated sequences (the RIP phenomenon of Selker et al.) was studied by tetrad analysis using ectopic copies of am+ (coding for NADP-specific glutamate dehydrogenase) and a missense allele am3, coding for a distinctive form of the enzyme, at the normal locus. In duplication crosses either both gene copies were inactivated or neither. Two inactivated am3 derivatives were shown to have undergone methylation and numerous base-pair changes, reflected in losses and gains of restriction sites, but without sequence rearrangement. Cutting at restriction sites within the disrupted sequences was incomplete but became almost complete following growth in the presence of 5-azacytidine. In a triplication cross in which one parent carried two unlinked ectopic gene copies together with am3 at the normal locus, premeiotic inactivation, when it occurred, tended to affect two of the three copies in any one ascus, but there were a few asci in which all three were inactivated.

Maintenance and differentiation in culture of pluripotential embryonic cell lines from pig blastocysts.

Cell lines were established from explanted blastocysts of domestic pigs; the cells could be maintained indefinitely when grown on mouse fibroblast feeder cell layers. They differentiate spontaneously at high densities, or when allowed to form aggregates when cultured on a non-adhesive substratum. Their appearance and differentiative behaviour resembles that of mouse embryonic stem cell lines. We are currently attempting to establish whether these cultures represent primary ectodermal lineages which would be of particular relevance to developmental and transgenic studies.

Derivation of pluripotent, embryonic cell lines from the pig and sheep.

As previously described for the establishment of stable, pluripotent cell lines from pig blastocysts, an analogous cell line was isolated from a sheep blastocyst. There are common features in the morphologies and growth characteristics of the pig and sheep cells in culture; in particular, pig and sheep cells display large nuclei and relatively sparse cytoplasm, as is observed in mouse embryonic stem cells. Furthermore, the morphology of the sheep cells closely resembles that of cells in primary cultures of inner cell masses isolated immunosurgically from sheep blastocysts. This suggests that the sheep cell line represents a primary ectodermal lineage.

[Role of computerized tomography following transrectal air insufflation and hypotonization and transrectal ultrasonography in the staging of rectal tumors]. / Ruolo della Tomografia Computerizzata eseguita dopo insufflazione di aria per via transrettale e ipotonizzazione e dell'ecotomografia transrettale nella stadiazione locale del tumore del retto.

Computed Tomography (CT) with rectal air inflation was compared with transrectal ultrasound (TRUS) in the preoperative staging of lower rectal cancer in 126 patients. Precontrast and postcontrast CT scans were performed with 5 mm thick slices; the rectum was previously inflated with air and antiperistaltic agents were administered. Preoperative results were compared with histologic findings. The accuracy, sensitivity and specificity of CT in predicting perirectal spread were 76%, 62% and 83%, whereas the corresponding figures for TRUS were 84%, 69% and 92%. The accuracy, sensitivity and specificity of CT and TRUS for nodal involvement were 58%, 60%, 57% and 72%, 68% and 66%, respectively. These results show that TRUS predicts perirectal spread and detects nodal metastases better than CT. However CT, when performed appropriately, shows tumor spread into perirectal fat and locoregional lymph nodes with high accuracy. Lymphatic involvement is strictly correlated with tumor size: TRUS and CT correctly staged only 57% and 43%, respectively, of the cases with nodal metastases and max. diameter of 5 mm. TRUS sometimes overstaged perirectal tumor growth (13 patients in our series) due to perirectal inflammation (9 cases) or artifacts caused by the presence of air bubbles between the probe and the tumor surface (4 patients). TRUS is a very useful tool for detecting tumor distance from the anal opening; in our series, the distance was incorrectly calculated only in one case (3 cm with TRUS versus 4 cm at surgery).