key: cord-0987170-lqcz1na6 authors: Vigara, LA; Villanego, F; Aguilera, A; García, T; Atienza, L; Pérez, J; García, A; Minguez, C; Montero, ME; Mazuecos, A title: Acute kidney injury, urinary and histopathological disorders in kidney transplant patients with SARS-CoV2 infection date: 2022-04-11 journal: Transplant Proc DOI: 10.1016/j.transproceed.2022.04.008 sha: aeb4a6e4e21c2e81b4a5f84e977df043f47658dd doc_id: 987170 cord_uid: lqcz1na6 INTRODUCTION Acute renal injury (AKI) is a manifestation of SARS-CoV-2 infection. The evidence in kidney transplants (KT) is limited. Specific, there are scarce data about the histological features in graft biopsies of these patients. MATERIAL AND METHODS A retrospective cohort study of KTs with SARS-CoV-2 infection from August 28, 2020 to April 23, 2021. We collected the incidence of AKI and the presence of urinary and histopathological disorders. Both groups were compared (AKI vs. no AKI). Immunohistochemical (IQ) and reverse transcription-polymerase chain reaction (RT-PCR) studies were performed on the anatomopathological samples. RESULTS Seventy-seven KT had SARS-CoV-2 infection. Twenty-seven patients (35.1%) developed AKI and it was related to increased severity and a worse evolution of the infection, defined by a greater presence of pneumonia (p <.001), hospitalization (p<.001), admission to the intensive care unit (ICU) (p<.001), and the need for ventilation support (p<.001) and continuous renal replacement therapy (CRRT) (p<.001). In the multivariable analysis, pneumonia behaved as an independent predictor for AKI development (p=.046). No differences were observed between proteinuria one month before and after infection (p=.224). Five patients showed microhematuria and two patients presented transient glycosuria without hyperglycemia. One of the five kidney biopsies performed (20%) showed positive SARS-CoV2 RT-PCR. CONCLUSION AKI is a frequent and potentially serious complication in KT patients. Occasionally it could be accompanied by abnormalities in the urinary sediment. One of the five biopsied patients had positive RT-PCR in renal tissue, which suggests the systemic spread of the virus and the tropism for the renal graft. Kidney involvement in Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection is common and its clinical presentation may range from mild proteinuria to acute kidney injury (AKI). Although the first works from China reported a low incidence of AKI in COVID-19 patients (3-9%) [1] , the current evidence shows that AKI rates are higher, reaching up to 43% in the rest of the series [2] . Furthermore, the incidence of AKI is markedly greater in critical patients since severe COVID-19 is likely to cause multi-organ failure [3] . In this sense, up to 79% of critical patients may develop AKI and some cases need renal replacement therapy (RRT). Thus, AKI is also considered a marker of severity and a negative prognostic factor for survival [2, 4] . In addition, some studies have described changes in urinary sediment in patients with SARS-CoV-2 infection, such as proteinuria, hematuria, and glycosuria, suggesting the presence of a renal reservoir for the virus [5] [6] [7] . Notwithstanding, kidney biopsies are not routinely performed in these patients, so there is no specific histological data. The information regarding AKI incidence and urinary abnormalities in kidney transplant (KT) recipients with SARS-CoV-2 infection is limited. However, these patients present several factors that can cause kidney impairment in addition to a higher risk of critical COVID-19 due to chronic immunosuppression and coexisting comorbidities. Therefore, specific studies in this population are necessary. Herein, we describe the incidence of AKI and the urinary and histopathological abnormalities as well as the risk factors for AKI in a cohort of KT recipients with SARS-CoV-2 infection. We performed a retrospective cohort study of all KT patients in our center with SARS-CoV-2 infection from August 28, 2020 (first case) to April 23, 2021. In all cases, the diagnosis was made by reverse transcription-polymerase chain reaction (RT-PCR) test in a nasopharyngeal swab. We defined AKI according to the Kidney Disease: Improving Global Outcomes (KDIGO) classification [8] . Clinical and analytical variables were collected, and patients with AKI were compared with those who did not develop AKI. We estimated glomerular filtration using the Modification of Diet in Renal Disease equation (MDRD-4) [9] . Patients were followed until recovery, defined by negative RT-PCR and/or IgG positive serology or until death. We also analyzed changes in the urine analysis during the infection (proteinuria, hematuria, or glycosuria). Proteinuria was determined by the albumin/creatinine ratio (ACR) in a first-morning urine sample. ACR the month before infection and one month later were compared. KT biopsies were performed if there was impaired renal function and/or urinary abnormalities such as previously described. In our center, samples of the upper pole of the kidney graft are taken by expert nephrologists using the 16-gauge biopsy under ultrasound-guide. All kidney biopsies were processed by standard techniques for light microscopy and immunofluorescence. Renal tissues were fixed in 10% neutral buffered formalin and embedded in paraffin. The processed sections were 5 µm each. Immunohistochemical studies were performed to detect SARS-CoV-2 with SARS CoV spike antibody, Rabbit Pab (IgG) and SARS CoV Nucleoprotein antibody, Mouse Mab (IgG), from Sino Biological laboratory. These antibodies are reactive on routine paraffin-embedded histological sections, using the avidin-biotin complex technique and heat-induced epitope retrieval (Optiview DAB IHC v6, Window). Immunohistochemical staining was performed with a 1:100 dilution for the nucleoprotein and 1:300 for the spike in an automated system (Ventana Benchmark ultra, Tucson, AZ). Heat-induced epitope retrieval was performed (4 minutes at 95°C for nucleoprotein and 8 minutes for spike), and the sample was subsequently incubated for 32 minutes for nucleoprotein and spike at room temperature. The histological samples were analyzed using the Applied Biosystems™ TaqMan™ Continuous variables are presented as mean and standard deviation or median and interquartile range as appropriate, and categorical variables as absolute value and percentage. The normality of the samples was analyzed by the Kolmogorov Smirnov test. All categorical variables were compared using Fisher exact test or Chi-square test. Continuous variables were compared using the Student's ttest or U Mann-Whitney for independent groups. In the case of paired groups, the Student's t-test for paired data was used or the Wilcoxon signed-rank test according to the normality of the sample. A multivariable logistic regression analysis was performed for AKI risk factors after SARS-CoV-2 infection. We used SPSS V.26.0 software (IBM, Armonk, NY, United States) for statistical analysis. In the study period, 77 KT had SARS-CoV-2 infection in our center. The mean age was 54.2 years, and 57.1% were men. Six KT recipients were in the first year post-KT and the median follow-up after infection was 20 days. Twenty-seven patients (35.1%) developed AKI. According to the severity of the AKI, they were classified as follows: AKI-I, n=13 (48.1%); AKI-II, n=6 (22.2%) and AKI-III, n=8 (29.6%). We compared baseline characteristics between groups (AKI vs. no AKI) ( Table 1) . Patients with AKI mostly had diabetes (p=.044) and had lymphopenia more frequently (p<.001). The presence of AKI was related to higher severity of the infection, defined by a greater presence of pneumonia (74.1% vs. 10%; p <.001), hospitalization (88.9% vs. 20% p <.001), admission to the ICU (40.7% vs. 2%; p <.001), and the need for ventilatory support (37% vs. 2%; p<.001). We found no differences in age, sex, hypertension, immunosuppressive treatment, baseline renal function, or time post-KT. The incidence of AKI in inpatients was 70.5% (n=24) and in critical patients (n=12), it reached 91.7% (n=11) and its was more severe, defined by a higher proportion of patients with AKI-III (54.5% vs. 12.5%; p=.033) and RRT (45.4% vs. 0%; p=.006). The mortality rate in patients with AKI was higher (51.9% vs. 0%; p<.001), and those with AKI III had a mortality rate higher (75%) than in patients with AKI I (30.8%) and AKI II (66.7%). All patients with AKI who required RRT (n=5) died. Patients cured of COVID-19 recovered baseline renal function. In the multivariable analysis, pneumonia was an independent predictor for AKI in KT patients with SARS-CoV-2 infection (p=.046) ( Table 2) . Regarding urinary abnormalities, we did not observe any changes in ACR between the value one month before infection and one month later ( Renal biopsy was performed in 5 patients, and 4 of them were recent KT ( Table 3 ). The median time post-KT to biopsy was 12 [6, 515] days. In three patients, the biopsy was performed due to impaired renal function, showing one of them, chronic toxicity due to anticalcineurin agents. In contrast, the rest did not show significant alterations. The fourth patient was a very highly-sensitized KT patient (98% PRA), whose renal biopsy showed an acute antibody-mediated rejection. The last case was a recent KT with non-normalization of renal function. Renal biopsy presented acute tubular necrosis (ATN), and the RT-PCR test in the anatomopathological sample was positive for SARS-CoV 2. SARS-CoV-2 IgG antibodies against spike and nucleoprotein in anatomopathological samples were negative in all renal biopsies performed. We present one of the first studies regarding AKI and urinary abnormalities after SARS-CoV-2 infection in a cohort of KT patients. Furthermore, we have confirmed the presence of viral genes in the histopathological samples. In our study, the AKI rate in inpatients with COVID-19 was 70.5%, reaching up to 91.7% in critical patients. This rate is markedly higher than in the general population, where the incidence ranges from 27-43% and 19-78% respectively [2] . Mortality in KT patients with AKI is also higher than in the series currently published in the non-KT population [10] . These data highlight the greater severity and the poor outcomes of COVID-19 in KT patients. Patients with AKI had a higher hospitalization rate, increased ICU admission and ventilator support. Besides, they needed continuous RRT and developed pneumonia more frequently, which is consistent with other series reports in both KT patients [4] and the general population. Due to this greater severity, AKI determines a bad prognosis of the infection, especially in hospitalized patients [11, 12] . In addition, AKI has been associated with higher mortality in patients with COVID 19, especially in more severe cases of AKI. An US study reported a mortality rate of 35% in patients with AKI; of those who died, 91% had AKI-III. The fatality rate was 55% in those needing RRT [8] . Besides,another study showed an in-hospital mortality rate of 33.7% in those with COVID-19associated AKI compared with 13.4% in those with AKI without COVID-19. Patients with AKI-III and COVID-19 had a 2.6-fold higher mortality rate than those with stage AKI-III who did not have COVID-19 [13] . In our study, half of patients with AKI died and those with AKI-III had a fatality rate higher than in patients with AKI 1 or 2. In our series, all patients with AKI who needed RRT died. These results show that renal impairment in KT patients with covid is related to worse outcomes that in general population, reflecting the greater vulnerability of KT recipients to the infection. Hirsh et al. reported that age and diabetes mellitus were risk factors for developing AKI in patients with SARS-CoV-2 infection, [11] . In our study, we have also documented a greater rate of diabetes in KT recipients with AKI. Although these patients were older, age was not statistically significant, probably due to a higher incidence of COVID-19 in our cohort of elderly KT patients. Additionally, patients with AKI presented more frequently lymphopenia (p<.001), similar to reported by other series of KT patients with COVID-19 [2, 12] . In our work, pneumonia was an independent predictor for the development of AKI in KT patients with COVID-19. The fundamental pathophysiology of pneumonia in critically ill patients is severe acute respiratory distress syndrome and the development of a multi-organ failure, which have been identified as independent risk factors for AKI [3, 14] . Pei. G et al. described the severity of pneumonia as a negative independent predictive factor of kidney complications in non-KT patients. [15] . Regarding alterations in the urinary sediment, hematuria and proteinuria after COVID-19 have been described in several studies [4, 16] . Notwithstanding, in our series we found no differences between albuminuria in the month before and after infection. We did not include albuminuria values at the time of infection due to the absence of data, especially in critical patients, where their clinical situation did not allow us to quantify it. Consequently, we can not determine if there was transient proteinuria. Hematuria was present in 5 patients and glycosuria without hyperglycemia was present in 2 patients. These latest discoveries could be related to glomerular and tubular damage of the virus, as other authors have already elucidated [5] [6] [7] . In this study, we have also described the histological findings in KT patients with COVID-19. without finding specific alterations. The most prevalent finding in these biopsies was ATN (62%) [17] . Kudose et al. evaluated 17 biopsy samples of native and allograft kidneys from patients with COVID-19. The most frequent findings in these biopsies were ATN (29%) and collapsing glomerulopathy (29%) [16] . In our biopsied KT patients, 1 of them, a recent KT without recovery of renal function, had ATN as the main finding. Notwithstanding, he also presented a positive RT-PCR for SARS-CoV-2 in renal tissue. These findings suggest that COVID-19 can spread through the bloodstream and infect other organs such as the kidney graft. Immunohistochemistry (IQ) failed to detect the virus in all kidney tissue samples analyzed as in other studies [18, 19] . Our results could show the greater sensitivity of RT-PCR with respect to IQ for the detection of SARS-CoV-2 in histological samples of renal tissue. We cannot clearly show the influence that time after infection can have on the negativity of both tests. Our study presents several limitations. This is a retrospective, single-center study with the limitations that inherently may exist in data collection. Therefore, we want to emphasize the exploratory nature of this study and we hope the findings will serve as a base for future studies. Finally, we do not have histological samples from patients with a fatal evolution of SARS-CoV-2 infection that probably reflects better the histological damage caused by this virus. Additionally, we do not have proteinuria values at the time of infection for the reason described. In conclusion, AKI is a frequent and potentially serious complication in KT patients that causes high hospitalization rates and ICU admission, mechanical ventilation, CRRT, and pneumonia. Occasionally, it could be accompanied by abnormalities in the urinary sediment. One of the five biopsied patients had positive RT-PCR in renal tissue, which suggests the systemic spread of the virus through the bloodstream and the tropism for the renal graft. 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