Properties of the feline tumour suppressor reduced expression in immortalized cells (REIC/Dkk-3).

Reduced expression in immortalized cells (REIC/Dkk-3), a member of the human Dickkopf (Dkk) family, is a growth suppressor in human and canine mammary tumours. Mammary gland tumours are common neoplasms with high malignancy in female cats. The purpose of this study was to clone the feline REIC/Dkk-3 homolog, investigate its expression in cell lines established from feline mammary gland tumours, and test its tumour suppressor function. Western blot analysis revealed that expression of the REIC/Dkk-3 protein was reduced in feline mammary carcinoma cell lines. Forced expression of REIC/Dkk-3 induced apoptosis in feline mammary tumour cell lines. These results demonstrate that REIC/Dkk-3 expression, which is downregulated in feline mammary tumour cell lines, results in the induction of apoptosis in these cells. Our findings suggest that feline REIC/Dkk-3 represents a potential molecular target for the development of therapies against feline mammary cancers.

Effects of Uric Acid on the NO Production of HUVECs and its Restoration by Urate Lowering Agents.

BACKGROUND: Although urate impaired the endothelial function, its underlying mechanism remains unknown. We hypothesized that urate impaired nitric oxide (NO) production in human umbilical vein endothelial cells (HUVECs) via activation of uric acid transporters (UATs). PURPOSE AND METHOD: In the present study, we studied effects of urate on NO production and eNOS protein expression in HUVEC cells in the presence and absence of urate lowering agents using molecular biological and biochemical assays. RESULTS: HUVECs expressed the 4 kinds of UATs, URATv1, ABCG2, MRP4 and MCT9. Exposure to urate at 7 mg/dl for 24 h significantly reduced production of NO. Pretreatment with benzbromarone, losartan or irbesartan normalized NO production. The same exposure resulted in dephosphorylation of endothelial NO synthase (eNOS) in HUVECs. Again pretreatment with benzbromarone, losartan or irbesartan abolished this effect. CONCLUSION: Urate reduced NO production by impaired phosphorylation of eNOS in HUVEC via activation of UATs, which could be normalized by urate lowering agents.

Ethmoidal sinusitis caused by Exserohilum rostratum in a patient with malignant lymphoma after non-myeloablative allogeneic peripheral blood stem cell transplantation.

We describe a patient with aggressive lymphoma who contracted an ethmoidal sinus infection due to Exserohilum rostratum after non-myeloablative allogeneic peripheral blood stem cell transplantation. E. rostratum is an extremely rare causative pathogen of invasive fungal infection. Phylogenetic tree analysis of the D1/D2 domains within the LSU rDNA identified the molecular structure of isolates. We believe this is the first description of E. rostratum infection in a patient who underwent hematopoietic stem cell transplantation.

Intravascular malignant lymphomatosis: diffusion-weighted magnetic resonance imaging characteristics.

Intravascular malignant lymphomatosis is an unusual condition in which malignant lymphoma cells form microscopic masses within the blood vessels of the central nervous system. Occlusion of the involved blood vessels can lead to multifocal cerebral infarcts. Diffusion-weighted magnetic resonance imaging (MRI) reveals a subacute infarction pattern (bright high signal intensity on b = 1000 s/mm2 images and intermediate apparent diffusion coefficient values) in the cerebral deep white matter. We present MRI findings of a 68-year-old woman with intravascular malignant lymphomatosis involving the cerebral white matter and the thoracic cord.

Epitope analysis of myeloperoxidase-specific antineutrophil cytoplasmic autoantibodies (MPO-ANCA) in childhood onset Graves' disease treated with propylthiouracil.

AIM: This study aimed to elucidate the relationship between epitope profiles and clinical manifestations of patients with myeloperoxidase antineutrophil cytoplasmic autoantibodies-(MPO-ANCA) positive childhood onset Graves' disease treated with propylthiouracil (PTU). METHODS: Sixteen patients were studied. The patients were grouped into ten without clinical vasculitis and nephritis (non-vasculitis group) and six with biopsy-proven pauci-immune necrotizing crescentic glomerulonephritis (vasculitis group). Epitope analysis was performed on serum samples by an enzyme-linked immunosorbent assay (ELISA) using a panel of recombinant deletion mutants of MPO. RESULTS: The high frequency sites were region upstream of Met341 (Ha region) near the N-terminus of the heavy chain, and regions downstream of Gly598 (Hf and Hg regions) near the C-terminus. Most patients in the non-vasculitis group had polyclonal MPO-ANCA recognizing both the above linear sites and other epitope sites of the heavy chain of MPO. Only one of ten patients in the non-vasculitis group, and four of six patients in the vasculitis group had MPO-ANCA recognizing only the linear sites of the heavy chain of the MPO molecule (Ha, Hf and/or Hg). Of the four patients in the vasculitis group, two had nephritis, like rapidly progressive glomerulonephritis and one had alveolar hemorrhage. CONCLUSION: These findings suggest that most patients with childhood onset Graves' disease treated with PTU who manifest no vasculitis have polyclonal MPO-ANCA recognizing both the linear and other epitope sites of the heavy chain of MPO. However, some patients who develop nephritis have MPO-ANCA recognizing only the linear sites of the heavy chain of MPO. This clonality of MPO-ANCA may be a risk factor that induces clinical vasculitis and nephritis in patients treated with PTU. Therefore, patients exposed to PTU should be monitored for MPO-ANCA level and epitopes.

[Report on the ECR2003 (European Congress of Radiology): non-contrast intracranial MR venography with 3D-phase contrast imaging: optimization of presaturation pulse and velocity encoding].

BACKGROUND: Intracranial MR venography is useful for the diagnosis of dural sinus thrombosis and the preoperative assessment of sinus patency encased by tumors. Recently, contrast-enhanced MR venography has been applied for suspected dural sinus occlusion in a shorter time. However, it has some disadvantage for the evaluation of hypervascularized enhancing thrombus mimicking flow in chronic sinus thrombosis. So far, we have evaluated optimal imaging technique and slice orientation and have shown that sagittal three-dimensional (3D) -phase contrast (PC) imaging is the most suitable for the non-contrast intracranial MR venography. PURPOSE: To assess the optimal presaturation pulse (SAT) and velocity encoding (VENC) for the non-contrast intracranial 3D-PC MR venography. METHODS AND MATERIALS: Firstly, we performed phantom experiment to assess the best SAT thickness using arterial presaturation. Second, MR imaging was performed in 7 healthy volunteers to measure the dural sinus flow velocity using a 1.5 T MR. Third, 3D-PC MR venography was performed with a VENC settings at 10, 15, 20 and 30 cm/sec for healthy volunteers. All data were displayed as maximum intensity projection images and three neuroradiologists assessed the visibility of the dural sinuses and the cortical vein. RESULTS: The mean flow velocity of the dural sinuses was 6.3 cm/sec. The thickness of the best SAT was 100 mm. In the assessment of the visibility of the 3D-PC images, dural sinuses were adequately visualized at a VENC of 15 cm/sec. CONCLUSIONS: Non-contrast intracranial 3D-PC MR venography was optimized at 100mm thickness of SAT and a VENC of 15 cm/sec.

The effect of cage stiffness on the rate of lumbar interbody fusion: an in vivo model using poly(l-lactic Acid) and titanium cages.

STUDY DESIGN: A goat interbody fusion model using poly-(L-lactic acid) and titanium cages was designed to evaluate the effect of cage stiffness on lumbar interbody fusion. OBJECTIVE: To investigate the effect of cage stiffness on the rate of interbody fusion. SUMMARY OF BACKGROUND DATA: Various types of cages considerably exceed the stiffness of vertebral bone, which ultimately may lead to postoperative complications. To avoid these complications, poly-(L-lactic acid) cages with limited stiffness have been designed. The mechanical integrity of the cages remains intact for at least 6 months. METHODS: Interbody fusions were performed at L3-L4 of 15 Dutch milk goats, and one of three cages was randomly implanted: 1) a titanium cage (n = 3), 2) a stiff poly-(L-lactic acid) cage (n = 6), or 3) a flexible poly-(L-lactic acid) cage (n = 6). Interbody fusion was assessed radiographically by three independent observers 3 and 6 months after surgery. RESULTS: At 3 months, all the poly-(L-lactic acid) specimens showed ingrowth of new bone, but with radiolucency in the fusion mass. At 6 months, solid arthrodesis was observed in four of six poly-(L-lactic acid) specimens, advanced ingrowth in one specimen, and infection in one specimen. Titanium cages showed ingrowth of bone, but with radiolucency in the fusion mass. Interbody fusion using poly-(L-lactic acid) cages showed a significantly higher rate statistically (P = 0.016) and more complete fusion than titanium cages of the same design. CONCLUSIONS: The reduced stiffness of poly-(L-lactic acid) cages showed enhanced interbody fusion, as compared with titanium cages after 6 months. Bioabsorbable poly-(L-lactic acid) cages thus may be a viable alternative to current interbody cage devices, thereby avoiding the concomitant problems related to their excessive stiffness. However, the bioabsorbability of the poly-(L-lactic acid) cages awaits investigation in a long-term study currently underway.

Clinicopathological study of unilateral multiple breast cancer.

BACKGROUND: The tendency for breast cancer to form multiple lesions is important to consider when planning breast-conserving surgery. However, many unknowns remain regarding the pathology and prognosis of multiple breast cancer, and therefore it is clinically significant to investigate its clinicopathological properties. METHODS: Over the past 25 years, in the period between April 1972 and March 1997, we investigated the clinicopathological findings including the 5-year and 10-year survival rates of 66 patients treated for unilateral multiple breast cancer. RESULTS: Of the total of 1,334 female patients with unilateral breast cancer who underwent curative surgery at our hospital, we identified 66 (5.0%) patients with unilateral multiple cancer. The incidence of such cancer has been higher in recent years. Of the 66 patients, 50 (75.8%) were premenopausal, and the remaining patients were postmenopausal, but multiple cancer among postmenopausal women is a recent phenomenon. The ER positivity rate of the main lesion in patients with multiple breast cancer was 69.2% and that of PgR was 50.0%. The 5-and 10-year overall survival rate in all 66 patients with multiple breast cancer was 90.8% and 79.7%, respectively. CONCLUSION: In the past, multiple breast cancer was frequently identified in premenopausal women. However, the current findings indicate that its incidence among postmenopausal women has increased in recent years. In addition, prognoses were comparable for patients with multiple or solitary breast cancer, a relevant finding in the planning of breast-conserving surgery.

Development of acute myeloid leukemia after chemotherapy for osteosarcoma.

The current study describes two patients with osteosarcoma who had acute myeloid leukemia develop after treatment with multiagent chemotherapy. The incidence density for the chemotherapy protocol was 129.8 per 10,000 person-year of followup. Karyotype analysis of 16 reported patients (including the current two patients) indicated that most leukemias after treatment of osteosarcoma correlated with the use of topoisomerase II inhibitors, such as doxorubicin. The deoxyribonucleic acid-damaging activity of doxorubicin reinforced by the use of alkylating agents is highly suspected as a causative event in the development of leukemia after treatment of osteosarcoma. As the next step in the development of treatment for patients with osteosarcoma, the type and intensity of treatment must be evaluated to minimize possible leukemogenic effects without compromising the potential for cure.

Comparison of various techniques of iron oxide-enhanced breath-hold MR imaging of hepatocellular carcinoma.

The purpose of our study is to compare qualitatively and quantitatively the abilities of various superparamagnetic iron oxide (SPIO)-enhanced breath-hold magnetic resonance imaging (MRI) techniques to detect hepatocellular carcinoma (HCC). Eight patients with HCCs were imaged. The images were obtained with conventional T2-weighted spin-echo imaging (CSE), half-Fourier single-shot turbo spin-echo (HASTE), single-shot gradient-echo type echo planar imaging (GE-EPI), and single-shot spin-echo type echo planar imaging (SE-EPI) before and after SPIO administration. The liver signal-to-noise ratios (SNRs) in CSE and each EPI sequence were significantly decreased after SPIO administration. GE-EPI had the highest decrease ratio (DR) of liver SNR, followed by SE-EPI (TE=98), SE-EPI (TE=28), CSE, and HASTE in this order. The relative contrasts with GE-EPI and SE-EPI (TE=98) were significantly higher than that with CSE after SPIO administration. On receiver operating characteristic (ROC) analysis, diagnostic accuracy did not differ significantly among the pulse sequences after SPIO administration. GE-EPI and SE-EPI (longer TE) were useful for SPIO-enhanced breath-hold MRI performed to detect HCC.

A novel alpha-amino-acid esterase from Bacillus mycoides capable of forming peptides of DD- and DL-configurations.

A novel alpha-amino-acid esterase possessing some properties favorable for the synthesis of D-amino acid-containing peptides has been purified from the culture broth of Bacillus mycoides. The enzyme consisted of 4 subunits of 39 kDa, had an isoelectric point of 7.0, and showed its maximum activity at around 47 degrees C and pH 7.6. The enzyme activity was strongly depressed by phenylmethanesulfonyl fluoride, but not by penicillin G or ampicillin, suggesting that the protein is a serine enzyme lacking penicillin-binding ability. The enzyme hydrolyzed a variety of D- and L-amino acid methyl esters with concomitant formation of homooligomers from D-Phe, D-Trp, D-Tyr, and D-Asp(OCH(3)) methyl esters, but it did not act on the D- or L-amino acid amides tested. Incubation of a mixture of Ac-D-Phe-OMe and D-/L-Leu-NH(2) with the enzyme yielded Ac-D-Phe-D-/L-Leu-NH(2) together with Ac-D-Phe-OH, the hydrolysate of the carboxyl component. To its credit, the enzyme failed to hydrolyze casein as well as peptides including diastereomers of diphenylalanine and dialanine, indicating that the enzyme would not cause secondary hydrolysis of once-formed peptides. These observations indicate the potential utility of the newly isolated enzyme for the synthesis of D-amino acid-containing peptides.

Reconstruction of the proximal humerus with the clavicle after tumor resection: a case report.

Reconstruction of the proximal humerus after resection for tumor and modification of the clavicular transposition procedure is described in which the blood supply of the clavicle is preserved and the clavicle is used to bridge the defect. An 11-year-old boy presented with shoulder pain, and the diagnosis was osteosarcoma of the right proximal humerus. After resection of the sarcomatous proximal humerus, the clavicle was released with its periosteum remaining intact, and the clavicle was rotated downward around the acromioclavicular joint. A vascularized fibula supplemented the reconstruction in trying to gain length of the arm. The acromioclavicular joint and the vascular supply of the clavicle were preserved. Internal fixation from the clavicle and the fibula to the distal humerus was made with an AO plate and screws. Muscles around the proximal humerus were reattached to the clavicle. Range of motion of the shoulder was 80 degrees flexion, 85 degrees abduction, 30 degrees external rotation, and 90 degrees internal rotation. Although the postoperative followup is relatively short, only 2 years, the functional advantages of this operation over other forms of reconstruction can be observed.

Cytotoxic T lymphocyte antigen 4 gene polymorphism confers susceptibility to type 1 diabetes in Japanese children: analysis of association with HLA genotypes and autoantibodies.

OBJECTIVE: Although the polymorphisms of the cytotoxic T lymphocyte antigen 4 (CTLA4) gene have been shown to be associated with Type 1 diabetes in Caucasians, some conflicting results have been reported among subjects of different ethnic backgrounds. We examined a CTLA4 polymorphism and its relationship to human leucocyte antigen (HLA) genotypes and autoantibodies for glutamic acid decarboxylase 65 (GAD65) and IA-2 in Japanese children with Type 1 diabetes. SUBJECTS AND MEASUREMENTS: The study group consisted of 125 childhood-onset Japanese subjects (50 males, 75 females) with Type 1 diabetes. The CTLA4 A/G polymorphism at position 49 was analysed using a PCR-restriction fragment length polymorphism (PCR-RFLP) method. The HLA-DRB1 and DQB1 genotypes were defined by DNA analysis using PCR-sequence-specific oligonucleotide (PCR-SSO) probes. The GAD65 autoantibody (GAD65Ab) and IA-2 autoantibody (IA-2Ab) titres were measured using radioimmunoassay. RESULTS: The distribution of genotype frequencies differs between subjects with Type 1 diabetes (GG: 46%, AG: 50%, AA: 5%) and controls (GG: 39%, AG: 44%, AA: 17%) (P < 0.01). The frequency of the G allele is higher in the diabetes group than in the controls (P < 0.05). When the subjects were subdivided according to HLA genotype, the two major HLA high-risk groups, with DR9-DQ9 and DR4-DQ4, that are unique to Japanese populations showed no difference in their CTLA4 polymorphism frequencies. Although no association between the CTLA4 polymorphism and the prevalence of GAD65Ab was found, CTLA4 GG subjects that had been newly diagnosed (< 9 months) had significantly higher levels of autoantibodies than AG subjects (P < 0.01). The prevalence and titres of IA-2Ab were not associated with the CTLA4 polymorphism. CONCLUSIONS: The CTLA4 gene might confer a susceptibility to childhood-onset Type 1 diabetes in the Japanese population. The association between this CTLA4 polymorphism and the HLA genotype was similar for both major groups with HLA high-risk alleles. CTLA4 might contribute to the humoral immune response to GAD in newly diagnosed subjects.

Angioimmunoblastic T-cell lymphoma with hyperplastic germinal centers: a clinicopathological and immunohistochemical study of 10 cases.

An absence of germinal centers is one of the histological characteristics of angioimmunoblastic T-cell lymphoma (AITL). We report here 10 unusual cases of AITL with hyperplastic germinal centers. The clinical presentation of each patient was characterized by generalized lymphadenopathy, constitutional symptoms and polyclonal hypergammaglobulinemia. The initial biopsy findings of each patient were similar and were characterized by hyperplastic germinal centers with ill-defined borders and a proliferation of high endothelial venules (HEV). In the paracortical area there was a mixed infiltrate including irregularly shaped clusters or small nests of clear cells in all cases. Moreover, the clear cells invaded the lymphoid follicles, resulting in expansion of the germinal centers, except for one case. Immunohistochemistry revealed that the tumor cells, including clear cells, were CD4-expressing T cells. Some of the atypical lymphocytes were also Bcl-6-positive. A majority of the follicular dendritic cell networks showed a normal/reactive or an expanded/disrupted pattern in all cases. Moreover, three lesions possessed a few large irregularly shaped proliferations of follicular dendritic cells around the HEV Four cases progressed to AITL within a few years. The present 10 cases probably represent an early stage of AITL preceding follicular dendritic cell hyperplasia. Detection of clear cells, Bcl-6-positive atypical T lymphocytes, and foci of irregularly shaped proliferation of follicular dendritic cells appears to be critical for early diagnosis and treatment of AITL with hyperplastic follicles.

[A case report of the patient at a terminal stage of stomach cancer, who had a visiting nurse].

We have started the visiting nurse system in our Kimitsu Central Hospital since September 1990, and have currently had 33 patients with 4 visiting nurses as of February 2001. Out of these 33 patients with the visiting nurse system, 9 patients are at a terminal stage of diseases. In the present study, we reported here a case of a 63 year-old female patient, who was at a terminal stage of stomach cancer and wished to have a regular home life, having a visiting nurse at home. The patient was not informed of her diagnosis as a cancer, but was convinced of her symptoms by her primary care physician that she was under a good care. Meanwhile, the patient showed less pain than that she was expected to have as a patient at the terminal stage of cancer. A daughter of the patient (actual caretaker) was first considering of hospitalization as an inpatient. However, since the patient strongly desired to stay and get care at home, the caretaker started a dual life of taking care of her mother at home and of raising children in parallel, in aware of a case of pronouncing death at home. While the patient's condition was relatively stable, the good quality of life (QOL) was maintained as she could make a trip of one-night stay. As the symptom progressed, however, the caretaker had a high anxiety with a pressure of accepting her own mother's death. On the other hand, a visiting nurse, who understood the patient's strong desire to stay at home and live her life as usual, also assisted in reducing the caretaker's high anxiety. Therefore, the visiting nurse basically assumed one possibility that the patient would die at home, but also prepared for the another possibility that the caretaker could choose to send her mother back to hospital whenever she felt her limit for being a caretaker. Moreover, the visiting nurse supported the patient and her family with an encouragement that they could live their favorable life till the end. As a result, the caretaker could continue to keep the patient at home, although she had a tremendous fear about taking care of her at home. When the patient's condition suddenly changed and got worsened, however, the caretaker desired to have her mother stay and get medicated in the hospital. Thus, the patient was sent to our hospital by ambulance. Although the caretaker did not have her mother die at home under her direct care, she recognized the great advantages in having a visiting nurse, who really assisted them well. Thus, it was confirmed that participation of a visiting nurse in the home healthcare could give the patient and her family a better, satisfactory life till the end. In general, although the patients or their family members wish to stay home and to have a home healthcare service, their desire or mind can be easily changed according to their situation. Especially in a case of the patient at a terminal stage, the family can be easily influenced by the condition of progression of the disease. One of the major roles of visiting nurses is always to understand the feelings of the patients and family and to prepare for the best as they can.

Total sacrectomy and reconstruction: oncologic and functional outcome.

The oncologic and functional outcomes of nine patients who were treated by total sacrectomy through L5 (three cases) or L5-S1 (six cases) were reviewed. Histologic diagnoses were one osteosarcoma, two giant cell tumors, two chondrosarcomas, and four chordomas. Patients' ages ranged from 17 to 70 years (mean age, 44.5 years). Resection margins were intralesional (giant cell tumors) in two, marginal in one, and wide in six patients (one contaminated). Reconstruction was performed using polymethylmethacrylate in two, screw and plate fixation in one, and a custom-made device in one. In five patients no reconstruction was performed. Five patients (45.5%) had wound complications: one had a wound dehiscence and two had deep infection; all needed surgical reintervention. In addition, in one a ventral and in another a dorsal hernia developed; only the ventral hernia was revised successfully. One patient had a deep vein thrombosis that was treated with a Coumadin derivate. Three patients (33%) died after 14, 18, and 50 months postoperatively respectively. One died of lung and widespread metastases, and two died of local recurrence and metastases. One patient with a giant cell tumor had a solitary lung metastasis. After resection the patient has been disease-free more than 90 months. At followup, six patients had no evidence of disease (mean followup, 73 months; range, 30-120 months). Functionally, there was no correlation between patients who had a reconstruction and those who had not. Total sacrectomy is a valuable procedure to secure local tumor control and overall survival, despite potential complications and neurologic and sexual dysfunction.

[A noteworthy case of postoperative liver metastasis from gastric cancer which responded well to UFT therapy].

A 66-year-old male underwent partial gastrectomy (resection of the pyloric side of the stomach) at another hospital on the basis of a diagnosis of gastric cancer (Fig. 1). Five months later, his CEA level began to rise. At that time, multiple liver metastases were detected by ultrasonography and CT scans. The patient received oral UFT therapy (400 mg/day) at our hospital. A reduction in CEA was observed 63 days after the start of this therapy. A judgment of CR (complete response) was made after 4 months of the therapy. At present, 2 years and 4 months after UFT was first administered, the patient shows no signs of tumor recurrence. This case is noteworthy since there has been no previous report of a case where UFT showed a high efficacy in treating liver metastasis after surgical resection of gastric cancer.

Pathologic examination of renal cell cancer by means of step-sectioning.

BACKGROUND: Because incidental detection of small renal cell cancers (RCC) has recently become increasingly common, nephron-sparing surgery for these cases has been more widely performed. Renal cell cancer was investigated by means of step-sectioning in order to determine which cases were suitable for nephron-sparing surgery and how it should be performed. METHODS: Pathology specimens obtained from 90 cases of radical nephrectomy were examined in 5 mm sections. We excluded large and invasive tumors from the present study. Particular attention was given to satellite tumor nodules (STN), pseudocapsules of the main tumor and vascular invasion. RESULTS: Satellite tumor nodules of cancer (STN-Ca) were found in seven of 90 cases (7.8%). Tumor size and grade was not correlated with the existence of STN-Ca. The distance between the main tumor and STN-Ca varied. with the pathology of the STN-Ca mostly resembling that of the main tumor. Pseudocapsules of the main tumor were incomplete in 53 cases (58.9%), but extracapsular invasion of more than 1 mm was not seen in tumors less than 50 mm in diameter. Through step-sectioning, a further six cases with microscopic vascular invasion were found. Vascular invasion within the main tumor was confirmed in 16 of 17 cases (94.1%) and within normal renal tissue in only one case. CONCLUSIONS: The incidence of existing STN-Ca was not high (7.8%) in the present study, we had not any characteristics in the cases with STN-Ca. As STN-Ca were rarely near the main tumor, we could not expect to resect STN-Ca with main tumor, resection to more than 1 mm outside the pseudocapsule was needed for complete resection of main tumor less than 50 mm in diameter. Normal renal tissue between the tumor and pelvis is a requirement for selecting appropriate cases in nephron-sparing surgery, but we could not remove the risk of leaving STN-Ca completely.