1. Clinical Presentation

Early recognition of graft dysfunction, rejection, and graft-versus-host disease (GVHD) manifestations is critical. Presentations vary significantly by organ type and timing, broadly categorized as acute versus chronic processes.

A. Solid Organ Rejection Signs

• Kidney: Increased serum creatinine, oliguria, graft tenderness, low-grade fever, and new-onset or worsening hypertension.
• Liver: A cholestatic pattern of liver function tests (elevated alkaline phosphatase and bilirubin), right upper quadrant pain, or general discomfort.
• Heart: Decreased exercise tolerance, new symptoms of heart failure (dyspnea on exertion, orthopnea), arrhythmias, or new conduction delays on ECG.
• Lung: Dyspnea, cough, hypoxemia, and new infiltrates on chest imaging. Chronic rejection often manifests as progressive, irreversible airflow obstruction, known as bronchiolitis obliterans syndrome (BOS).

B. Graft-versus-Host Disease (GVHD) Manifestations

Unique to hematopoietic stem cell transplant (HSCT) recipients, GVHD occurs when donor immune cells attack the recipient’s tissues.

• Acute (typically ≤100 days post-transplant): Presents with a classic triad of a maculopapular rash, anorexia/nausea with high-volume diarrhea (>500 mL/day), and elevated bilirubin.
• Chronic (typically >100 days post-transplant): A more diverse, systemic syndrome with scleroderma-like skin changes, painful oral ulcers, dry eyes (keratoconjunctivitis sicca), cholestatic liver injury, and bronchiolitis obliterans.

2. Laboratory Diagnostics

Laboratory analysis is essential for differentiating rejection from other common complications like infection or drug toxicity. The approach combines routine chemistry, therapeutic drug monitoring, infection surveillance, and emerging biomarkers.

Histopathology: The Gold Standard

Despite advances in non-invasive testing, tissue biopsy remains the definitive method for diagnosing and grading rejection.

• Kidney: The Banff criteria are used to score interstitial inflammation (i-score), tubulitis (t-score), and vascular lesions to diagnose acute cellular rejection.
• Heart: The ISHLT grading system (0R to 3R) is based on the presence and extent of lymphocytic infiltrates and associated myocyte injury.
• GVHD: Biopsies of affected tissues (skin, gut, liver) showing characteristic epithelial apoptosis and lymphocytic infiltration confirm the diagnosis.

3. Imaging and Invasive Modalities

Imaging is used to rule out anatomic or vascular complications and to guide invasive procedures. Endoscopic and transbronchial sampling provide the tissue necessary for a definitive histologic diagnosis.

• Ultrasound with Doppler: First-line imaging for most solid organ transplants. In renal transplant, it assesses graft perfusion via resistive indices and can identify hydronephrosis or fluid collections (hematoma, lymphocele). In liver transplant, it confirms patency of the hepatic artery, portal vein, and biliary tree.
• CT/MRI: Provide detailed anatomical information to evaluate for vascular thrombosis, anastomotic leaks, infarcts, or parenchymal changes in the lungs (e.g., ground-glass opacities, air trapping).
• Echocardiography: Standard for monitoring cardiac allograft function, assessing for new wall motion abnormalities, quantifying ventricular function, and detecting pericardial effusions.
• Endoscopic and Transbronchial Biopsies: The cornerstone of diagnosis for GI GVHD (via upper endoscopy/colonoscopy) and lung rejection (via bronchoscopy with transbronchial biopsies).

Immunopathology

Specialized staining and antibody testing are critical for diagnosing antibody-mediated rejection (AMR), a distinct and often more aggressive form of rejection.

• C4d Staining: C4d is a complement split product that deposits in peritubular capillaries during AMR. A positive C4d stain on a biopsy specimen is a key diagnostic criterion.
• Donor-Specific Antibodies (DSA): The presence of circulating antibodies in the recipient’s serum that target the donor’s HLA antigens is the hallmark of AMR.

4. Classification and Severity Scoring

Standardized scoring systems are vital for communicating severity, determining prognosis, and triggering appropriate treatment intensification.

Other Key Scoring Systems

• ISHLT Grades (Heart): 0R (no rejection), 1R (mild), 2R (moderate, focal myocyte injury requiring treatment), and 3R (severe, diffuse injury with high risk of hemodynamic compromise).
• ISHLT A/B System (Lung): Grades acute rejection (A0-A4) and airway inflammation/lymphocytic bronchiolitis (B0-B2R) separately.
• NIH Chronic GVHD Staging: Assigns organ-specific scores (0-3) across multiple systems (skin, liver, lung, etc.) to calculate a global severity score (mild, moderate, or severe).

5. Clinical Algorithms

Stepwise diagnostic and therapeutic pathways are essential for standardizing care, ensuring timely evaluation, and guiding treatment decisions for urgent conditions like acute rejection and GVHD.

Suspected Acute GVHD Pathway

1. Initial Assessment: Quantify and stage the rash percentage, daily diarrhea volume, and total bilirubin level.
2. Grading: Assign an overall grade from I (mild) to IV (severe).
3. Treatment Initiation: For patients with Grade II-IV acute GVHD, initiate systemic corticosteroids (e.g., methylprednisolone 1-2 mg/kg/day).
4. Response Assessment: Re-evaluate clinical status at day 7. Patients who do not show significant improvement are considered steroid-refractory.
5. Second-Line Therapy: For steroid-refractory GVHD, add a second-line agent such as a calcineurin inhibitor, ruxolitinib, or another approved therapy.

6. Controversies and Emerging Tools

The field of transplant diagnostics is rapidly evolving, with a major focus on developing non-invasive tools to reduce the need for biopsies and to personalize patient care.

Non-invasive Tests Under Investigation

While biopsy remains the gold standard, several “liquid biopsy” technologies are gaining traction:

• Donor-derived cell-free DNA (dd-cfDNA): Already in clinical use, these panels measure fragments of donor DNA in the recipient’s blood as a marker of graft injury. High sensitivity makes it an excellent screening tool, but specificity can be affected by other forms of graft injury besides rejection.
• Gene Expression Profiling (GEP): Measures the expression of a panel of genes in peripheral blood mononuclear cells to create an “immune quiescence” or “rejection” signature.
• Immune Cell Phenotyping: Uses flow cytometry to analyze populations of immune cells (e.g., regulatory T-cells, effector memory T-cells) to predict rejection risk.

Standardization Challenges and Future Directions

A major hurdle is the variability in diagnostic practices between transplant centers. This includes differences in biopsy scoring by pathologists, different thresholds for laboratory assays, and inconsistent application of staging systems.

The future lies in combining multiple data points. Machine learning algorithms are being developed to integrate clinical data, trough levels, dd-cfDNA trends, and GEP scores into a single, comprehensive risk score. Centralized digital pathology review and large international registries are essential for validating these new tools and harmonizing practices worldwide.