case_images: PMC11371627_01
case_images: Overview of XK pathophysiology and clinical manifestation in our patient. (A) The XK gene codes for the XK protein which consists of ten transmembrane domains and forms a heterodimer with the Kell protein via a disulfide bond (Cys347
  XK - Cys72
  Kell). The XK protein is expressed in many tissues, including RBCs, bone marrow, brain and other neuronal tissues. The KEL gene codes for the Kell protein, which is a glycoprotein of 732 amino acids. The Kell protein is expressed on RBCs, bone marrow, testes and fetal liver. The protein expresses 25 antigens, the best known Kell blood group antigens are Kell (K) and Cellano (k) located at amino acid position 193, Kp at amino acid position 281 and Js at amino acid position 597. (B) Photograph. The patient's lower legs with dominant atrophy of the left gastrocnemius muscle. (C) Whole body muscle MRI. Bilateral fatty oedematous atrophy of adductors, hamstring muscles and calves with T1 hyperintensity (red arrow). Hyperintensity in STIR sequences revealed recent oedema in the above muscles (light grey arrowhead). Muscles in other parts of the body were not affected. (D) Histological examination after biopsy of the left gastrocnemius muscle with hematoxylin and eosin staining, myosin slow and myosin fast staining revealed severe atrophy of skeletal muscle with vacant fat (black arrow) and surplus of connective tissue, but without signs of inflammation, rimmed vacuoles or ragged red fibers. No other specific structural or immunohistochemical abnormalities were detected. Abbreviations: Cys, cysteine; MRI, Magnetic resonance imaging; F, front; H/E, hematoxylin eosin staining; R, righthand side; RBCs, red blood cells; STIR, Short Tau Inversion Recovery sequence.
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case_images: F1
case_images: Figure 1B shows difference in calf circumference with a slimmer left lower leg and atrophy of the gastrocnemius muscle. MRI showed bilateral fatty and oedematous atrophy of adductors, hamstrings and both calves (Figure 1C). MRI of the lumbar spine showed multisegmental degeneration with spinal narrowing at L3/4 and L4/5 but no significant neural foraminal constrictions or myelopathy (Supplementary Figure S1A). No other specific structural or immunohistochemical abnormalities were detected (Figure 1D). For an overview of clinical work-up of our patient, please see Supplementary Figure 1B.
case_images: CSF flow cytometry analysis, molecular genetic analysis and KEL gene expression analysis in our patient. (A) CSF flow cytometry. Flow cytometry analysis of the patient's CSF revealed elevation of HLA-DR+CD8+ T cells with a CD4/CD8 ratio of 1.4 and elevation of NKs. (B) Sanger sequencing of the XK gene revealed a novel deletion at position 756, resulting in a frameshift deletion and a premature termination of the amino acid chain (exon 3; c.756del, p.Trp253Glyfs*15). Segregation analysis in the mother revealed that they carry the same XK deletion. (C) Fluorescence-activated Cell Sorting was performed to analyse the expression of Kell antigens Kell (K) and Cellano (k) on erythrocytes. Five samples of fresh EDTA and two samples of RBCs were detected with antibodies (a) anti-k and (b) anti-K. Our patient is marked in red. (c) Presentation of the Kell antigens Cellano (k) and Kell (K) in our patient. (D)
  KEL gene expression assay. Real-time PCR was performed using the TaqMan Gene Expression Assay. Our patient with the novel XK deletion (F) showed no restriction in expression of the KEL gene on RNA level. Abbreviations: CD56bright NK, CD56bright natural killer cells; CD56dim NK, CD56dim natural killer cells; cMono, classical monocytes; CSF, cerebrospinal fluid; del, deletion; EDTA, Ethylenediaminetetraacetic acid; Gly, glycine; HLA-DR pos, activated cells; iMono, intermediate monocytes; ncMono, non-classical monocytes; NK, natural killer cells; NKT, natural killer T cells; PCR, polymerase chain reaction; RBCs, red blood cells; RNA, ribonucleic acid; Trp, tryptophane.
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case_images: PMC11371627_01_fgene-15-1421952-g002.jpg
case_images: F2
case_images: CSF standard analysis showed no abnormalities in total cell count or protein and no intrathecal immunoglobulin synthesis. CSF flow cytometry revealed a slight elevation of HLA-DR CD4+ (27.8%, reference <24%) and HLA-DR CD8+ cells (75.5%, reference <63,4%), a CD4/CD8 ratio of 1.4 and an elevation of natural killer cells (NKs) (5,8%) (Figure 2A). Multi-gene panel sequencing including common candidate genes for myopathy or muscular dystrophy revealed a novel hemizygous deletion (c.756del, p.Trp253Glyfs*15) of the XK gene (Figure 2B). In order to examine its impact, we performed FACS analysis of blood which revealed negativity for Kell (K) and a reduced density of Cellano (k) antigen on the patient's erythrocytes in comparison to healthy controls (Figure 2C). Real-time PCR revealed no restriction of KEL gene expression in our patient (Figure 2D).