[Hemostasis disorders in patients with systemic AL-amyloidosis].

AIM: To analyze the frequency and nature of hemorrhagic and thrombotic complications in patients with systemic AL-amyloidosis and compare with laboratory changes in the hemostasis system. MATERIALS AND METHODS: The prospective study included 40 patients with newly diagnosed AL-amyloidosis. To detect amyloid, all patients underwent bone marrow trephine biopsy and duodenal biopsy, and 28 (70%) patients underwent biopsy of the affected organ. Before the start of therapy, all patients were determined the platelet count, activated partial thromboplastin time, thrombin time, fibrinogen concentration, time of XIIa-dependent fibrinolysis, antithrombin III, D-dimer, activity of blood coagulation factors VIII, X and vWF. The statistical part of the study was carried out using the IBM SPSS Statistics 2017 system software (SPSS, Chicago, IL, USA). RESULTS: In 20 (50%) patients, hemorrhages on the skin and mucous membranes were diagnosed as vascular purpura. Before the start of therapy, 7 (17.5%) patients had thrombosis, including leg vein thrombosis (5 patients), ischemic stroke (2 patients). There was a direct correlation between thrombotic complications and cutaneous hemorrhagic syndrome (p=0.007). In 15 (75%) cases, cutaneous hemorrhagic syndrome was accompanied by hypercoagulable shifts in the hemostasis system. Of the 20 patients with cutaneous hemorrhagic syndrome, 19 (95%) patients had kidney damage, including 15 patients with nephrotic syndrome. Hematoma type of bleeding, as well as heavy bleeding was not observed, including after a biopsy of the internal organs. According to the totality of hemostasis indicators, hypercoagulation syndrome was more often observed (in 23; 56% of patients). Hypocoagulation was diagnosed only in 2 (5%) patients with liver damage, 16 (39%) patients had normocoagulation. CONCLUSION: Cutaneous hemorrhagic syndrome is the most common clinical manifestation of disorders in the hemostasis system in patients with AL-amyloidosis. The relationship of hemorrhages on the skin with nephrotic syndrome has been established, which may indicate a single pathogenetic mechanism. Cutaneous hemorrhagic syndrome is associated with hypercoagulable shifts in hemostasis and a high risk of thrombotic complications.

[Detection of paraprotein in plasma cell tumors].

Paraprotein is a laboratory biomarker of plasma cell tumors and other lymphoproliferative diseases. Its determination is necessary for diagnosing, monitoring and assessment of therapy effectiveness. The lecture presents the main methods of qualitative and quantative analysis of monoclonal proteins: gel electrophoresis, capillary electrophoresis, immunofixation and nephelometry features, possibilities and limitations are reviewed. The main sources of errors and artifacts during these studies are considered. Also the difficulties in the diagnosis and interpretation of the results of serum and urine tests are highlighted.

[Morphological and immunohistochemical predictors of renal response to therapy patients with myeloma cast nephropathy and dialysis-dependent acute kidney injury].

AIM: Reveal morphological and immunohistochemical predictors of reversibility of dialysis-dependent acute kidney injury (AKI) in patients with myeloma cast nephropathy (MCN) based on the study of kidney biopsy. MATERIALS AND METHODS: Renal pathological findings were studied in 36 patients with MCN and dialysis-dependent stage 3 AKI (AKIN, 2012). The study of biopsy samples was performed by a semi-quantitative and quantitative analysis using computer morphometry. The expression of E-cadherin, vimentin and-smooth muscle actin was determined immunohistochemically in the tubular cells and interstitium. Induction therapy for 26 patients was carried out to bortezomib-based programs; in 10 patients other schemes were used. A comparative analysis of morphological changes in nephrobiopathy depending on the renal response was performed in patients with achieved hematologic remission. RESULTS: Improved renal function was observed only in patients with hematologic response to therapy. There were no differences in the number of sclerotic glomeruli, protein casts, the area of inflammatory interstitial infiltration, and the degree of acute tubular damage in patients with and without renal response. In patients with renal response compared with patients without improving renal function, the area of interstitial fibrosis was less (24.9% and 45.9%, respectively;p=0.001), and the area of E-cadherin expression was larger (15.9% and 7.1%, respectively;p=0.006). Interstitial fibrosis of 40% or more and/or the area of expression of E-cadherin less than 10% of the area of tubulo-interstitium have an unfavorable prognostic value in achieving a renal response in MCN. CONCLUSION: If the interstitial fibrosis area is 40% or more and the expression area of E-cadherin is less than 10%, the probability of the absence of a renal response is 93.3% (OR=24.5) even when a hematological response to induction therapy is achieved. The number of protein casts, the prevalence of acute tubular damage and inflammatory interstitial infiltration have not prognostic value.

[Bone disease as the first manifestation of systemic AL-amyloidosis].

Our case demonstrates severe bone disease in primary AL-amyloidosis without concomitant multiple myeloma. A 30-year-old man had spontaneous vertebral fracture Th8. A computed tomography scan suggested multiple foci of lesions in all the bones. In bone marrow and resected rib werent detected any tumor cells. After 15 years from the beginning of the disease, nephrotic syndrome developed. Based on the kidney biopsy, AL-amyloidosis was confirmed. Amyloid was also detected in the bowel and bone marrow. On the indirect signs (thickening of the interventricular septum 16 mm and increased NT-proBNP 2200 pg/ml), a cardial involvement was confirmed. In the bone marrow (from three sites) was found 2.85% clonal plasma cells with immunophenotype СD138+, СD38dim, СD19-, СD117+, СD81-, СD27-, СD56-. FISH method revealed polysomy 5,9,15 in 3% of the nuclei. Serum free light chain Kappa 575 mg/l (/44.9) was detected. Multiple foci of destruction with increased metabolic activity (SUVmax 3.6) were visualized on PET-CT, and an surgical intervention biopsy was performed from two foci. The number of plasma cells from the destruction foci was 2.5%, and massive amyloid deposition was detected. On CT scan foci of lesions differed from bone lesions at multiple myeloma. Bone fragments of point and linear type (button sequestration) were visualized in most of the destruction foci. The content of the lesion was low density. There was no extraossal spread from large zones of destruction. There was also spontaneous scarring of the some lesions (without therapy). Thus, the diagnosis of multiple myeloma was excluded on the basis based on x-ray signs, of the duration of osteodestructive syndrome (15 years), the absence of plasma infiltration in the bone marrow, including from foci of bone destruction by open biopsy. This observation proves the possibility of damage to the skeleton due to amyloid deposition and justifies the need to include AL-amyloidosis in the spectrum of differential diagnosis of diseases that occur with osteodestructive syndrome.

[Autologous haematopoietic stem cell transplantation in patients with multiple myeloma complicated by dialysis-dependent renal failure].

AIM: To assess the safety and efficacy of autologous haematopoietic stem cell transplantation (auto-HSCT) in multiple myeloma (MM) patients with dialysis-dependent renal failure. MATERIALS AND METHODS: During a period from May 2010 to December 2016 fourteen MM patients with dialysis-dependent renal failure aged 48 to 65 years underwent auto-HSCT. After the induction therapy complete response, very good partial response, partial response were documented in 64, 29, 7% of patients, respectively. In no case was a renal response achieved. Haematopoietic stem cell mobilization in most patients (13/14) was performed according to the scheme: G-CSF 10 g/kg. Melphalan in 3 dosages was used as pre-transplant conditioning: 100, 140 and 200 mg/m2; 13 patients underwent a single and in one case underwent a tandem auto-HSCT against the background of hemodialysis. Evaluation of the antitumor and renal response was assessed on the 100th day after auto-HSCT. Subsequently, against the background of programmed hemodialysis and in the setting of high-dosed melphalan (100200 mg/m2), 13 patients underwent a single and one patient underwent a tandem auto-HSCT. At +100 days after auto-HSCT, an antitumor response and renal response were assessed. RESULTS: The period of agranulocytosis after auto-HSCT was from 5 to 12 days (median 8,5) and was accompanied by infectious complications, cardiac and neurological dysfunctions. At +100 days after auto-HSCT, the complete response was confirmed in 71% patients and very good partial response was confirmed in 29% patients. The minimal renal response was registered in 2 patients (14%), hemodialysis was stopped. The transplant-related mortality was absent. After a median follow-up of 53 months 5-year progression-free survival was 59%, and overall survival was 93%. CONCLUSION: Carrying out auto-HSCT in patients with dialysis-dependent renal failure contributed to the achievement of a minimal renal response in 14% of cases, which allowed these patients to stop hemodialysis. Patients whose conditioning regimen was performed using melphalan at a dose of 200 mg/m2showed more frequent complications in the early post-transplant period compared to patients who received a lower dose of melphalan (100140 mg/m2). Auto-HSCT in MM patients with dialysis-dependent renal failure is a feasible and effective treatment method, which in some cases contributes to independence from hemodialysis.

[Difficulties evaluating hematological response in patients with multiple myeloma and dialysis - dependent renal impairment].

AIM: to determine serum free light chains (FLC) level by patients with multiple myeloma (MM) and dialysis - dependent renal impairment in which the amount Bence Jones (BJ) protein in the urine met the criteria of hematological response. PATIENTS AND METHODS: This study included 13 MM with dialysis - dependent renal impairment patients (estimated glomerular filtration rate < 10 ml/min), whose urine BJ protein content was less than 200 mg/day after antimyeloma therapy (including 11 patients whose urine BJ protein content was less than 100 mg/day). RESULTS: The median serum concentration of monoclonal FLC was 608.7 (298-8380) mg/l. Thus, with trace amounts BJ protein in the urine serum content monoclonal FLC varied 28 times with the same degree of severity of renal failure. In patients with oliguria serum SLC content was significantly higher than in normal diuresis (1109 and 307 mg/L; p.

[Current approaches to treating of patients with multiple myeloma with renal failure: Questions and proofs]. / Sovremennye podkhody k lecheniiu bol'nykh mnozhestvennoi mielomoi s pochechnoi nedostatochnost'iu: voprosy i dokazatel'stva.

Renal failure (RF) is detected in 20-30% of patients at the onset of multiple myeloma (MM), in 50% of patients during its progression. The advent of new, highly effective agents has considerably expanded the possibilities of treatment in MM patients. Unfortunately, patients with RF, especially those with severe RF, were not included in the majority of investigations. The available data are based on the results of treatment in small groups of patients generally without the morphological identification of nephropathies, with varying severity of RF, which explains significant differences in renal response rates. This review analyzes the results of the most important studies and gives recommendations for treatment in accordance with national and international standards.

[Light-chain deposition disease is a hematologic problem]. / Bolezn' depozitov legkikh tsepei ­ gematologicheskaia problema.

AIM: To analyze clinical and laboratory data and treatment results in patients with light-chain deposition disease (LCDD). SUBJECTS AND METHODS: Nine patients with LCDD and kidney injury were examined. The diagnosis was based on the results of light and immunofluorescence microscopy of renal biopsy specimens. All the patients received bortezomib, cyclophosphamide, and dexamethasone (VCD) induction therapy. RESULTS: Six patients were diagnosed with multiple myeloma; in 3 patients LCDD was considered within monoclonal gammopathy manly involving the kidney. By the initiation of therapy, all the patients were diagnosed as having chronic kidney disease (Stage III (n=2), Stage IV (n=2), and dialysis-related renal failure (n=5)). After the VCD treatment, 7 of 9 patients achieved a hematologic response. Second-line therapy with lenalidomide proved to be effective in the other 2 cases. Five patients achieved complete remission; 3 had a very good partial remission. Thereafter, 2 patients received high-dose melphalan chemotherapy and autologous hematopoietic stem cell transplantation. Better renal function was noted in only 2 cases. CONCLUSION: Despite the high efficiency of therapy aimed to reduce monoclonal light chains; improved renal function was observed in only 2 (22%) patients. Such low rates of a renal response were due to the late initiation of therapy.

[Uromodulin gene polymorphisms in patients with cast nephropathy in multiple myeloma]. / Polimorfizmy gena uromodulina u bol'nykh kast-nefropatiei pri mnozhestvennoi mielome.

AIM: To investigate the nature of mutations in exons 4 and 5 of the uromodulin (UM) gene, including in the area encoding the domain of 8 cysteines (D8C), in patients with multiple myeloma (MM) with the secretion of monoclonal light chains (LC) in cast nephropathy (CN) and without kidney injury. SUBJECTS AND METHODS: The investigation enrolled 24 patients in MM remission, who were observed to have monoclonal LC secretion at onset. Group 1 included 14 patients with CN; Group 2 consisted of 10 patients with normal renal function (a comparison group). The compared groups did not differ in the number of serum and urinary monoclonal LCs. Genomic DNA was extracted from the peripheral blood samples of patients. The nucleotide sequence of exons 4 and 5 of the UM gene was determined by the Sanger method. RESULTS: No differences were found in the frequency of polymorphisms depending on the severity of kidney injury. The missense mutation p.142R>R/Q in the UM gene, which had not been previously described, was discovered. CONCLUSION: The patients with MM were not found to have statistically significant differences in the frequency and nature of polymorphisms of exons 4 and 5 in the UM gene, including in the area encoding D8C, in CN without kidney injury.

[Secondary monoclonal gammopathy after bone marrow autotransplantation as a cause of worse renal function in light chain immunoglobulin deposition disease].

The paper describes a clinical case of a female woman with nephropathy due to light chain deposition disease caused by secretion of κ Bence-Jones protein. Complete immunochemical remission was achieved after induction therapy using a bortezomib + cyclophosphamide + dexamethasone regimen. Renal function remained unchanged (glomerular filtration rate 16 ml/min), there was a reduction in proteinuria from 5.8 to 2.6 g/day. High-dose melphalan (200 mg/m2) chemotherapy with peripheral blood stem cell autotransplantation was performed as consolidation of remission. A year posttransplantation, there was no secretion of κ light chains; however, monoclonal IgG lambda emerged in a quantity of 3.2 g/l. At the same period, nephrotic syndrome became progressive (daily proteinuria 12 g) and dialysis-dependent renal failure developed. A repeat renal biopsy specimen revealed changes, suggesting that there was a decrease in renal deposits of κ light chains. Simultaneously with this, the obvious negative trend as progressive nephrosclerosis and fixation of IgG and λ light chains in the glomeruli (in the sclerotic areas) cause IgGλ monoclonal protein to be involved in the genesis of further kidney injury. Attention is also paid to different characteristics of capillary wall deposits by density (according to the electron microscopic findings), which may point to their different qualitative composition and possibly different formation duration. Papaprotein Gλ disappeared after a year without therapy, suggesting its reactivity. The findings confirm that worse renal function is caused by the action of paraprotein Gλ due to secondary (after autologous hematopoietic stem cells transplantation) monoclonal gammopathy.

[Oligosecretory monoclonal gammopathy with renal involvement]. / Oligosekretornaya monoklonal'naya gammapatiya s porazheniem pochek.

The article deals with the so-called monoclonal gammopathy of undetermined significance (MGUS), which is being actively explored in the world and has been recently investigated in Russia. It indicates the principles of identifying the phenotypes of MGUS and criteria for assessing the risk of its progression to cancer. There is an update on the possible involvement of monoclonal proteins in the pathogenesis of certain non-neoplastic kidney diseases, renal injuries in particular. The paper gives their classification and enumerates differential diagnostic techniques, including the Freelite method, a highly sensitive one to determine free light chains (FLC), prognostic criteria, and approaches to treating each separate form in relation to the phenotype of a monoclonal protein. The authors present their own data on detection rates for MGUS at a multidisciplinary hospital and a clinical case of MGUS-associated membranoproliferative glomerulonephritis, by justifying a treatment regimen containing bortezomib (velcade).

[Dialysis-dependent renal failure in patients with multiple myeloma: Reversibility factors].

AIM: To establish the reversibility factors of dialysis-dependent renal failure (RF) in patients with multiple myeloma (MM) treated according to bortezomib-containing programs. SUBJECTS AND METHODS: The efficiency of treatment according to bortezomib-containing programs was evaluated in 40 patients with first diagnosed MM and dialysis-dependent RF. Prior to treatment, 34 patients underwent needle renal biopsy. The early mortality rate was 5%. RESULTS: After treatment according to bortezomib-containing programs, 83% of the patients could achieve a hematologic response, including 66% who had complete and very good partial remission (CR and vgPR). A renal response (RenR) was observed in only 26% of the patients. RenR to antitumor therapy was found to be determined by the morphological variant of nephropathy. Improved kidney function was observed only in cast nephropathy (CN) and absent in other types of kidney injury. In CN, the rate of RenR depends on the degree of renal tubulointerstitial fibrosis at the initiation of treatment. Kidney function improved in 20% of the patients with disseminated tubulointerstitial fibrosis and in 57% of those with minimal fibrotic changes (p=0.04). To achieve RenR, it is important to have an early antitumor response in addition to baseline morphological changes in the nephrobiopsy specimen. When the number of monoclonal light chains (LC) in urine was reduced to 100 mg/day after 2 cycles of induction therapy, RenR was 55%; with a lower antitumor response it was as high as 28% (p=0.04). A LC decrease to the values of vgPR after 2 cycles of induction therapy was noted in only 32% of the patients; 44% of the patients achieved vgPR or CR after an average of 6 (3-13) therapy cycles. CONCLUSION: In MM patients with dialysis-dependent RF, RenR was caused by the morphological variant of kidney injury and by the degree of tubulointerstitial fibrosis at the therapy initiation, as well as by the rate at which monoclonal LCs reduced.

[The concurrence of light-chain deposition disease, AL-amyloidosis, and cast nephropathy in a patient with multiple myeloma].

Despite of the fact that their clinical manifestations are similar, AL-amyloidosis (AL-A) and light chain deposition disease (LCDD) are individual nosological entities in view of considerable differences in their pathogenesis and pathomorphology. The paper describes a rare case of the concurrence of LCDD and AL-A in a patient with multiple myeloma. Clinically, there was dialysis-dependent renal failure, flail leg syndrome, myocardiopathy, and rhabdomyolysis. At the disease onset, his nephrobiopsy specimen could diagnose LCDD and myeloma or cast nephropathy. The disease was characterized by an aggressive course. Despite the administration of innovative agents, the patient had a short-term remission and died from disease progression. Autopsy additionally revealed amyloid deposition in the heart and kidney. The development of AL-A in the presence of prior LCDD may reflect the progression of the tumor and the appearance of an additional subclone of plasma cells that produce amyloidogenic light chains. The uncommonness of this case is that renal amyloid was found in the tubular casts and absent in the glomeruli, which may be considered as a special form--tubular AL-amyloidosis.

[Crystalline histiocytosis].

The paper describes a case of diagnosis of the rare monoclonal secretion-associated disease crystalline histiocytosis with kidney and bone marrow involvement. The female patient with multiple myeloma (MM) was found to have intralysosomal crystals in the cells of the bone marrow (histiocytes, plasmocytes), kidneys proper (mesangiocytes, podocytes), and subsequently in those of a kidney graft. Lower secreted monoclonal IgG and ceased Bence-Jones protein secretion after MM chemotherapy were accompanied by improved and stabilized kidney graft function. However, a repeat morphological study of a renal biopsy specimen showed that the crystalline inclusions were preserved in the podocytes. By comparing the immunological and renal responses, it is reasonable to suggest that monoclonal IgG rather than Bence-Jones protein is of value in the pathogenesis of crystal formation.

[Fibrillary glomerulonephritis and immunotactoid nephropathy: the current view of the pathomorphological substrate of disease].

Fibrillary glomerulonephritis (FGN) and immunotactoid nephropathy (ITN) are diseases diagnosed only by electron microscopy. Until recently, information on the diseases has reached as reports on some cases. Much information, including the authors' observations, has been presently gathered so as there is a chance of attempting to pool and analyze it. It is quite obvious that investigators do not agree to evaluate FGN and ITN. Some authors are inclined to believe that these are one disease and propose to use the general term "fibrillary-immunotactoid nephropathy", we, like others, consider FGN and ITN as two different diseases. Our findings suggest that there are two diseases (FGN and ITN). We provide support for the data available in the literature on that the deposits are polyclonal in FGN and monoclonal in ITN. It is most likely that no sharp distinction can be made between FGN and ITN from the diameter of microtubules making up deposits (less or more than 30 nm); the procedure of their package and, to be sure, their chemical composition are of importance in establishing the diagnosis.

[Nephropathy caused by non-amyloid organized and granular deposits as multiple myeloma syndrome].

A rare variant of nephropathy in multiple myeloma (MM) is reported. Nephropathy is characterized basing on the study of nephrobiopsy with light, immunofluorescent and electron microscopy. A repeat biopsy of the kidney was made after achievement of a complete clinicohematological remission. A MM patient's nephrobiopsy in a picture of glomerulonephritis had 3 types of deposits: granular, irregular fibrils of 12 nm in diameter and microtubes organized in bunches 19 nm in diameter. Congo red test was negative, cryoglobulinemia was absent. Immunofluorescent test detected deposit of monoclonal IgG in the mesangium and glomerular basal membrane (GBM) corresponding to monoclonal type of monoclonal secretion. After treatment and achievement of remission, neither IgG no light chains were found in nephrobiopsy. Electron microscopy registered complete resorption of granular deposits and microtubes with formation of electron-transparent cavities. However, fibrils seen before treatment only in mesangium appeared in the above hollow cavities. The presence of such fibrils in the mesangium and GBM did not influence clinical picture of the disease. After achievement of remission the patient had no clinical and laboratory signs of nephropathy, only insignificant selective glomerular proteinurea was observed (0,5 g/l). Thus, granular deposits and microtubes contained paraprotein, they were completely resorbed after achievement of MM remission. Fibrils seem to have another genesis unrelated to monoclonal gammapathy.

[Multiple myeloma in renal transplant recipients].

Severe renal failure (RF) may be the first and only clinical manifestation of multiple myeloma (MM). Occasionally the disease remains long unrecognized and the patients receive renal function replacement therapy, including renal transplantation (RT). To treat MM in renal transplant recipients is a complex medical and ethical problem. The paper presents the authors' experience in treating 3 patients with MM diagnosed after RT and evolving transplant lesion. Various morphological types of grafted kidney lesion were detected. These included fibrillar glomerulonephritis, cast nephropathy, and the latter concurrent with light-chain deposition disease. RF most rapidly progressed in cast nephropathy. The natural history of the disease was unfavorable in all patients; VAD and PAD chemotherapy programs proved to be ineffective. It is concluded that RT should not be performed in patients with extended-stage MM due to the fact that there is a considerable risk for renal transplant lesion and severe infectious complications that may occur during chemotherapy. Blood and urine immunochemical studies should be conducted in all the patients who are to undergo RT.

[Renal monoclonal immunoglobuline deposition disease as a rare variant of renal damage in multiple myeloma].

A case of the immunoglobuline deposit disease diagnosis in MM patient having the symptoms of intensive proteinuria, macrohematuria and terminal renal failure is reported. The disease was diagnosed by the evidence from electron microscopy of the kidney and liver biopsies.

[Treatment and survival of multiple myeloma patients on programmed hemodialysis].

AIM: To analyse clinical picture of multiple myeloma (MM) and treatment results in MM patients on programmed dialysis (PD). MATERIALS AND METHODS: Case histories of 22 MM patients were analysed. They had a terminal stage of chronic renal failure (CRF) in the onset of the disease. Chemotherapy (CT) was performed in 20 patients (10 patients received VAD program, the other 10--melfalan). RESULTS: Early lethality was 28%. The patients died of septic complications. Neutropenia was observed significantly more frequently on melfalan treatment than on VAD therapy (9 and 2 patients, respectively; chi-square 5.6; p = 0.009). Survival median, excluding early lethality, was 16 months. Differences by therapy were not registered. Three patients on MP program survived more than 3 years. Function of the kidneys improved in 4 (20%) patients. Hemodialysis was avoided in 2 patients. Survival of patients with reestablished renal function was maximal (44 and 84 months). CONCLUSION: Standard CT for MM with terminal CRF is associated with high toxicity and frequent septic complications. Survival is better if renal function improves and HD discontinues. Reversibility of CRF at a terminal stage in MM does not depend on completeness of hematological response. Programs with melfalan CT provoke more frequent myelotoxic cytopenia, early lethality is higher but the number of longer survivals is more.

[Concerning the condition of the bone marrow in patients with multiple myeloma]. / O sostoianii kostnogo mozga pri mnozhestvennoi mielome.

The bone marrow of 48 patients with multiple myeloma was studied prior to therapy. A relationships was established between cluster formation rate, on the one hand, and the numbers of myelocaryocytes, erythrocytes and circulating immune complexes, on the other. In patients with stage III disease, the diminished cellularity of the marrow was due to decreased levels of granulocytes and normocytes. Moreover, those patients revealed fewer erythroclastic marrow clusters. Similar changes in the granulocytic component of hemopoiesis were present in those who died within earlier stages (20 months) after the beginning of therapy. It is suggested that diminished granulocyte pool alongside reduced cluster formation rate may play a role in infectious complication development.