This was a retrospective, single-centre study of all patients undergoing first heart transplantation in Gothenburg, Sweden, between January 1988 and December 2000. During this period, 283 transplantations were performed in 278 patients. Of these, 34 patients died within 30 days of transplantation and were excluded from the analysis, as were 17 children < 14 years and one patient for whom the record from 1988 could not be found. In 226 patients, CMV infection, disease and potential risk factors were evaluated during the first 12 months after transplantation. Angiographic signs of CAV were recorded for 10 years or until death. Cardiovascular risk-factors were monitored according to protocol. Statins was given as universal therapy after 1996. Follow-up of all patients was complete through 31 December 2000. No patients were lost to follow-up.
The induction therapy during 1988–1993 consisted of cyclosporine A (CsA) and from 1994 of anti-thymocyte globulin (ATG; 2.5 mg/kg/day administered intravenously before surgery and for 3 to 5 days afterwards). Daclizumab or 100 mg of prednisone was given in the event of ATG allergy. Methylprednisolone was administered at 500 mg intravenously before surgery and 500 mg intra-operative and then at 125 mg every 8 hours for 3 doses during the whole study period. The maintenance immunosuppression therapy consisted of standard triple therapy. CsA (5 to 8 mg/kg/day) was used to maintain serum CsA levels within range 200–350 ng/ml during the first year and from 100 to 200 ng/ml thereafter. Azathioprine (AZA) was administered at 2 mg/kg/day, and prednisone at 0.2 mg/kg/day reduced to 0.1 mg/kg/day orally. Since 1995, Tacrolimus (TAC) was an alternative to CsA, given at 0.075 mg/kg to maintain serum tacrolimus levels within range 10–15 ng/ml and, since 1997, AZA was replaced by mycophenolate mofetil (MMF) (2–3 g/day).
Detection of CMV infection
CMV infection was detected by serology (seroconversion post-transplant), viral culture, qualitative polymerase chain reaction (PCR) for CMV DNA, histopathology and immunohistochemistry (IHC) with CMV-specific antibodies from endomyocardial biopsies (EMB) or tissue biopsies from other organs. Clinical symptoms of CMV disease were also documented for 12 months after transplantation.
Definitions of CMV infection
CMV infection: CMV virus detected by viral culture or qualitative, PCR assay for CMV in any body fluid or tissue specimen. Seroconversion from CMV (seronegative to seropositive) was also regarded as CMV infection.
CMV infection was categorised as either asymptomatic CMV infection or CMV disease.
Asymptomatic CMV infection: Evidence of CMV infection but not fulfilling criteria for CMV disease.
CMV disease: Evidence of CMV infection with attributable symptoms in accordance with Ljungman et al. . CMV disease was categorized as tissue-invasive disease or CMV syndrome with fever, leucopoenia and/or thrombocytopenia.
Prophylaxis, treatment and monitoring of CMV
All patients during 1988 to 1991: No CMV prophylaxis was given. Patients were tested frequently. Seronegative patients received treatment with intravenous (i.v.) ganciclovir or foscavir for 14–21 days in the event of seroconversion or a positive viral culture for CMV.
High-risk group (D+/R−) during 1992 to 1997: Pre-emptive treatment was given, comprising monitoring with qualitative CMV PCR once weekly during the first three months post-transplantation. When CMV DNA was detected in serum, patients received treatment with i.v. ganciclovir for at least 10 days. During 1998–2000, universal prophylaxis was given with 1,000 mg of oral ganciclovir tid for 14 weeks.
Intermediate-risk group (R+) during 1992 to 2000: No prophylaxis was given. Qualitative CMV PCR was analyzed in serum when CMV disease was clinically suspected.
In D+/R−transplants, serological analyses were repeated once monthly for the first 4 months after HTx, then at 6, 9 and 12 months and thereafter annually and when infection was suspected during 1988 to 1998.
CMV disease was treated with 5 mg/kg of i.v. ganciclovir bid for 10–21 days. Asymptomatic CMV infection was treated in seronegative recipients. Ganciclovir dosing was adjusted for renal function. Patients who developed severe CMV pneumonitis (hypoxia) also received polyclonal immunoglobulin.
Diagnosis of CAV
All available coronary angiographic studies performed in the study cohort between the first and tenth year of follow up, or until death or re-transplantation, were retrospectively re-analyzed visually. Where coronary angiography was not performed in patients who were alive during the study period it was due to medical contraindications or patient refusal. CAV-free survival was defined as the time to CAV of any grade observed visually by angiography. CAV was graded according to Costanzo et al. (none, mild, moderate or severe) . The severity of CAV (none, mild, moderate, severe) was assessed throughout follow up. Donor-related coronary artery disease was defined as no stenosis = 0 or no significant stenosis = 1 from coronary angiographic studies performed before HTx.
Risk factor analysis
Data on potential risk factors for CAV were collected retrospectively, including recipient and donor characteristics, cold ischemic time, cardiovascular risk factors (hypertension, diabetes mellitus, smoking status before transplantation and complications (acute rejection (AR) episodes).
Surveillance endomyocardial biopsies for AR were standardized for all patients and graded according to the 2005 ISHLT classification as 1R, 2R or 3R . AR therapy consisted of 1,000 mg boluses of methylprednisolone for 3 consecutive days in cases of AR ≥ 2R. Severe cellular rejections were treated with 2.5 mg/kg/day of ATG for 3 days. Clinical relevant antibody-mediated rejection (AMR) was treated with plasmapheresis. The cumulative effect of acute cellular rejection was assessed by the total rejection score (TRS)  and defined as 0R = 0, 1R = 1, 2R = 2, 3R = 3. Severe TRS was defined as all AR ≥ 2R. The scores were normalized for the total number of biopsy specimens taken during the first 12 months after HTx (TRS or TRS ≥ 2R) in the individual patient.
This study was approved by the local Ethical Committee of Gothenburg (115–14) and by the medical director of the heart transplant department at the University Hospital, Gothenburg, Sweden. The data were recruited from the patients’ medical records and local registries according to standards by the Declaration of Helsinki.
Data were analyzed using SPSS version 20.0 (SPSS Inc, Chicago, IL, US). Continuous variables are presented as mean values ± standard deviations (SD) and categorical variables as percentages. The chi-square test was used to compare proportions and occurrences between groups. Confidence intervals (CI) were calculated using a normality approximation algorithm. Survival and CAV-free survival time was analyzed using the Kaplan–Meier procedure and statistical comparisons of survival distributions between different categories were made using the log rank test. Cox’s univariate and multivariate model was used to determine risk factors for events. Variables in the univariate model testing with a p value < 0.1 were included in the multivariate model. A p value < 0.05 was considered statistically significant.