Howell-Jolly Body-Like Neutrophil Inclusions Following Hematopoietic Stem Cell Transplantation: A Case Report

Introduction

In 1989, Barbara J. Bain first described the neutrophil inclusions that resemble Howell-Jolly bodies in patients treated with azathioprine. These inclusions were named for their similarity to the Howell-Jolly bodies found in erythrocytes [1]. Although clinically infrequent, their occurrence is not only linked to the severity of infections but can also correlate with specific treatment responses, especially within the framework of bone marrow transplantation and immunosuppressive therapy. These inclusions are observed in conditions where bone marrow function is impaired or the spleen’s filtering capacity is diminished, carrying significant implications for clinical practice.

This case study augments the existing literature [2–6] by offering a distinctive case – a 14-year-old boy who presented with Howell-Jolly body-like inclusions following a hematopoietic stem cell transplant. This uncommon occurrence is rare among post-transplant patients and is associated with complications from COVID-19 infection and gastrointestinal graft-versus-host disease (GVHD), highlighting its clinical significance and rarity. Our study underscores the appearance of these inclusions after bone marrow transplantation and subsequent immunosuppressive therapy, providing a comprehensive record of the patient’s medication history and disease progression.

Our case provides a detailed narrative of the patient’s journey from transplantation to disease progression, with a particular focus on the clinical implications of Howell-Jolly body-like inclusions, especially as they emerge in routine blood smears as a signature of dual effects from immunosuppressive drugs and viral infections following bone marrow transplantation. This phenomenon may predict a heightened risk of severe infections and more serious complications of GVHD, offering clinicians a crucial early warning signal.

By conducting an in-depth comparison with existing literature, we found that although Howell-Jolly body-like inclusions have been reported in patients undergoing various types of organ transplants, our case exhibits uniqueness in several key aspects. First, our patient underwent a hematopoietic stem cell transplant, which may have distinct effects on the morphology and function of neutrophils. Second, while SARS-CoV-2 infection and gastrointestinal GVHD have been reported in other transplant cases, our patient also developed mucormycosis, a rare and severe complication potentially related to immunosuppressive therapy.

Through this case, we aim to emphasize the clinical relevance of Howell-Jolly body-like inclusions and explore the importance of incorporating their presence into treatment strategies for effective patient management. Our study offers a new perspective on the emergence of these inclusions following hematopoietic stem cell transplantation and may uncover novel mechanisms of inclusion formation or new associations with specific treatments.

Case Report

TREATMENT STRATEGY:

Given the patient’s diagnosis and the aggressive nature of the disease, we opted for allogeneic hematopoietic stem cell transplantation as the primary treatment strategy. On February 9, 2023, the donor (the patient’s father) showed 5 out of 10 HLA loci matches and blood type compatibility (O-type donor to B-type recipient). After a comprehensive assessment, no significant contraindications to transplantation were identified, and a decision was made to proceed with a haploidentical hematopoietic stem cell transplantation. The treatment strategy consisted of 3 phases: conditioning regimen, stem cell infusion process, and post-transplant care.

CONDITIONING REGIMEN:

To prepare the patient for transplantation and reduce tumor burden, we used the following conditioning regimen: total marrow irradiation to eliminate residual cancer cells in the bone marrow; idarubicin for its immunosuppressive effect; regimen of cladribine, cytarabine, and granulocyte colony-stimulating factor, to further suppress tumor cells and stimulate the mobilization of normal hematopoietic stem cells; post-transplant cyclophosphamide to prevent GVHD. This regimen was designed to maximize the success rate of stem cell transplantation while minimizing complications.

STEM CELL INFUSION PROCESS:

On the day of transplantation (day 0), following the conditioning regimen, the patient received an infusion of third-party unrelated umbilical cord blood (female donor to male recipient, B-type blood compatible with B-type blood, with 8 out of 10 HLA loci matches). This was due to the lower immunogenicity and rapid availability of umbilical cord blood stem cells, which could provide the patient with immediate immune protection. The infusion of umbilical cord blood provided the patient with a temporary immune barrier while awaiting engraftment and recovery of the donor stem cells. The infusion process was uneventful, with no adverse reactions. After transplantation, the patient was admitted to a specialized post-transplant care unit for close monitoring and supportive treatment.

POST-TRANSPLANT CARE:

On day +1 after transplant, the patient was administered intravenous immunoglobulin and antiviral drugs to prevent infection; concurrently, on day +3, cyclophosphamide and cyclosporine were used to prevent GVHD. Additionally, we closely monitored the patient’s blood routine and biochemical indicators to promptly identify and manage potential complications. The patient achieved successful platelet engraftment on day +12 and leukocyte engraftment on day +21. On day +28, a bone marrow aspiration was performed. This preliminary assessment included detection of minimal residual disease in the bone marrow, genetic and chromosomal analysis, chimerism, and viral testing. However, due to the lack of fluorescence in situ hybridization testing, we were unable to accurately assess the patient’s post-transplant chimerism status.

POST-TRANSPLANT COMPLICATIONS:

On December 13, 2023, the patient developed pulmonary GVHD, and subsequently, on February 14, 2024, was readmitted due to SARS-CoV-2 infection-related diarrhea, concurrently developing gastrointestinal GVHD. For pulmonary GVHD, the patient was initially treated with cyclophosphamide and ruxolitinib. In the management of gastrointestinal GVHD, due to the patient’s intolerance to cyclosporine and tacrolimus, the treatment regimen was adjusted. On March 13, 2024, continuous low-dose tacrolimus infusion was initiated, and when the condition worsened, on March 21, 2024, the treatment was escalated to a combination therapy including mycophenolate mofetil and ruxolitinib.

LABORATORY FINDINGS:

During the patient’s readmission for SARS-CoV-2 infection-related diarrhea, a series of auxiliary tests were conducted, with results summarized in Table 1. Notably, routine blood smear analysis revealed 1 to 2 neutrophils containing 1 to 3 small, round, deeply stained cytoplasmic inclusions, morphologically similar to Howell-Jolly bodies in red blood cells (Figure 1). Although a bone marrow aspiration was not performed concurrently for direct comparison, the presence of Howell-Jolly body-like inclusions in peripheral blood smears observed in this patient may have been related to previous viral infections and ongoing immunosuppressive therapy. These treatments may have led to abnormalities in the maturation process of neutrophils, characterized by irregular nuclear segmentation and atypical patterns of chromatin condensation, ultimately forming separate inclusions detached from the main nucleus. These findings have significant clinical implications as they may reflect the patient’s persistently suppressed immune system and abnormal responses to infection and drug treatment. Furthermore, these abnormal neutrophil inclusions may be associated with the risk of developing gastrointestinal GVHD. To further explore this phenomenon, we utilized our hospital’s laboratory information system to analyze the timing, frequency, and tacrolimus usage associated with the appearance of Howell-Jolly body-like inclusions in post-transplant blood smears, with relevant data detailed in Table 2. This analysis helps us better understand the immune status and complication risks of post-transplant patients, thereby providing a basis for individualized treatment.

DISEASE PROGRESSION:

Given the patient’s persistent diarrhea showing no signs of improvement, coupled with inadequate control of gastrointestinal GVHD, we decided to intensify the anti-infective treatment regimen. On March 24, 2024, following extensive multidisciplinary team consultation, it was recommended that the patient undergo gastroscopy and colonoscopy. Gastroscopy revealed black, mossy substances attached to the mucosal surfaces of the esophagus, stomach, and duodenum, accompanied by bleeding. Colonoscopy identified multiple ulcerative lesions in the colon, with similar black mossy substances attached to the ulcers, which were not easily removed after washing. To confirm the diagnosis, a biopsy of the colonic mucosa was performed for histopathological examination, next-generation sequencing, and fungal immunofluorescence monitoring. Laboratory results indicated the presence of fungal hyphae in the mucosal attachments, which were broad, aseptate, and branched at right angles, suggesting mucormycosis. Based on the comprehensive findings, a diagnosis of extensive gastrointestinal mucormycosis was made. In response to this diagnosis, the patient’s anti-fungal treatment regimen was upgraded to the maximum dose, while continuing current medications and adding oral amphotericin B. While continuing tacrolimus, all other immunosuppressants were discontinued. However, on the second night of treatment, on March 24, 2024, the patient suddenly developed progressive dyspnea, accompanied by a decrease in oxygen saturation. An emergency chest CT scan revealed multiple pulmonary infiltrates, further supporting the diagnosis of mucormycosis. After a comprehensive discussion with the patient’s family about the severity and irreversible nature of the condition, the family understood and chose to sign for voluntary discharge.

This case highlights the importance of vigilant monitoring of patients following allogeneic hematopoietic stem cell transplantation. The series of complications encountered by the patient, including pulmonary and gastrointestinal GVHD, as well as the rare gastrointestinal mucormycosis, reflect the complexity of post-transplant management. Most notably, the discovery of uncommon neutrophil inclusions in the patient may have been associated with previous viral infections and long-term immunosuppressive therapy. The manifestation of these inclusions may suggest a persistently suppressed immune system and potential abnormal responses to infection and drug treatment, which has significant implications for clinical diagnosis.

Discussion

In this case report, we meticulously documented the rare scenario of a 14-year-old boy who was re-hospitalized following a hematopoietic stem cell transplantation due to SARS-CoV-2 infection and gastrointestinal GVHD. During the treatment course, we observed the emergence of neutrophil inclusions resembling Howell-Jolly bodies, a phenomenon not commonly seen in post-transplant patients. Through an in-depth comparison with existing literature [3,4,8], we found that although Howell-Jolly body-like inclusions have been reported in patients undergoing various types of organ transplants, our case exhibits uniqueness in several key aspects. First, our patient underwent a hematopoietic stem cell transplant, differing from the commonly reported liver, heart, and lung transplants in the literature, which may have distinct effects on the morphology and function of neutrophils. Second, while SARSCoV-2 infection and gastrointestinal GVHD have been reported in other transplant cases, our patient also developed mucormycosis, a rare and severe complication potentially related to immunosuppressive therapy. Additionally, the Howell-Jolly body-like inclusions observed in our patient may have been associated with the combined effects of SARS-CoV-2 infection and immunosuppressive therapy, which has not been thoroughly explored in existing literature. These findings underscore the importance of monitoring neutrophil morphology in post-transplant patients and suggest that when such inclusions are observed, potential infections and the impact of immunosuppressive therapy should be considered. Our case also highlights the importance of monitoring and timely diagnosing infections in post-transplant patients and the need to consider these factors when devising treatment strategies.

Morphologically, neutrophil inclusions must be distinguished from apoptotic bodies and Barr bodies [10,11]. Apoptotic bodies typically appear during the process of apoptosis, manifesting as fragmentation of the cell nucleus, while Barr bodies are drumstick-shaped structures connected to the cell nucleus. In our case, the observed inclusions differed from these structures, often being separate from the main cell nucleus and exhibiting the typical characteristics of Howell-Jolly bodies in terms of morphology.

Currently, the mechanism behind the formation of Howell-Jolly body-like inclusions in neutrophils is not fully understood. Based on an analysis of past case reports, we believe that the use of tacrolimus, an immunosuppressant, may be a primary cause for the emergence of Howell-Jolly body-like inclusions. The occurrence of Howell-Jolly body-like inclusions following the use of tacrolimus in organ transplant patients, as documented in existing literature, is summarized in Table 3. Tacrolimus inhibits calcineurin, blocking T-cell activation and IL-2 production, affecting the function of T-helper cells and other immune cells, thereby potentially disrupting the maturation process of neutrophils, leading to abnormal nuclear segmentation and chromatin condensation patterns, resulting in the formation of Howell-Jolly body-like inclusions. It may also be due to unusual hypersensitivity reactions caused by the drug, leading to morphological changes in neutrophils [9]. Additionally, the drug can interfere with DNA synthesis, further affecting the normal maturation of neutrophils [7]. Second, certain viral infections, as observed in the early stages of HIV infection and in patients with SARS-CoV-2 infection [6,12], are also associated with the appearance of Howell-Jolly bodies. Viral infections can affect the function of marrow cells or directly act on the maturation process of neutrophils, leading to the formation of Howell-Jolly body-like inclusions. This case involves a patient who underwent a hematopoietic stem cell transplant and subsequently developed gastrointestinal GVHD. After treatment with the anti-GVHD drug tacrolimus, Howell-Jolly body-like inclusions were observed in the patient’s peripheral blood.

This case demonstrates the observation of Howell-Jolly body-like inclusions in the peripheral blood smear of a patient following a hematopoietic stem cell transplant. Nevertheless, this study has several limitations: First, due to the involvement of only a single hematopoietic stem cell transplant case, it is currently not possible to determine the exact association between drug use and the emergence of Howell-Jolly body-like inclusions. Therefore, future studies should expand the sample size to further explore the impact of drugs on the formation of Howell-Jolly body-like inclusions; second, this study did not assess the relationship between the presence of Howell-Jolly body-like inclusions in peripheral blood smears and the severity of disease infection. However, in this case report, the appearance of these inclusions in the patient’s peripheral blood smears suggests that the patient may have been experiencing a severe infection; finally, the morphological changes of Howell-Jolly body-like inclusions in neutrophils are not pathognomonic of specific diseases and should be considered in the context of the overall clinical situation. In particular, tacrolimus, as an immunosuppressant widely used in post-transplant patients, may be associated with morphological changes in neutrophils. Future studies should consider drug use, viral infections, and other factors that may affect neutrophil morphology, to more comprehensively understand the mechanism behind the formation of Howell-Jolly body-like inclusions.

Conclusions

In this study, we meticulously reassessed the neutrophil inclusions observed in the peripheral blood of patients following hematopoietic stem cell transplantation. Given the complex treatment regimens and multiple factors involved in managing post-transplant patients who developed gastrointestinal GVHD, it was not feasible to establish a direct link between the inclusions and the disease state. However, morphological analysis strongly suggested that these inclusions were likely induced by the medications administered as part of the treatment protocol. Additionally, the emergence of these inclusions may have also indicated the presence of a severe infectious agent within the patient’s system. Our findings underscore the potential role of immunosuppressive drugs and viral infections in the genesis of these inclusions.