Short-Term Treatment for Immune-Mediated Acquired Lecithin–Cholesterol Acyltransferase Deficiency Restores the High-Density Lipoprotein Function: A Case Report

Abstract

Familial lecithin–cholesterol acyltransferase (LCAT) deficiency with a primary LCAT gene mutation results in various conditions, including corneal opacity, anemia, kidney disease, and low high-density lipoprotein (HDL) levels. In recent years, secondary LCAT deficiency with nearly identical symptoms has been identified as a rare case of immune-mediated acquired LCAT deficiency caused by LCAT autoantibodies. In limited cases, prednisolone treatment is required for severe conditions and has been shown to favorably modulate LCAT autoantibodies and restore LCAT activity, resulting in improved HDL-cholesterol (HDL-C) levels, renal dysfunction, and other complications. However, there is little detailed information regarding LCAT activity, lipid changes, and renal dysfunction after the initiation of prednisorone treatment. In the present study, in addition to the effects on LCAT activity, lipids, and proteinuria, we for the first time monitored the HDL cholesterol efflux capacity (CEC), an important anti-atherosclerotic HDL function, during the first month of treatment in a patient with this disease. We found that the LCAT activity, HDL-C concentration, and HDL CEC increased from undetectable or low values to normal ranges during this period, as did proteinuria. Specifically, the HDL CEC and LCAT activity recovered faster than the HDL-C levels. Based on these findings, the effects of prednisolone treatment on LCAT and HDL CEC activities prior to HDL-C levels suggest that normal HDL-C levels may not be essential as a treatment target in immune-mediated acquired LCAT deficiency patients who require treatment.

1.Introduction

Low high-density lipoprotein-cholesterol (HDL-C) levels are associated with coronary artery disease¹⁾. A recent large-scale general population study in Japan found that low HDL-C levels may predict future renal dysfunction²⁾. Lecithin–cholesterol acyltransferase (LCAT) promotes HDL maturation by increasing esterified cholesterol levels. Familial LCAT deficiency (FLD), caused by a primary gene mutation, results in various conditions, such as corneal opacity, anemia, kidney disease, and low HDL levels³⁾. FLD is partially linked to the development of atherosclerosis³⁾, as well as the presence of low HDL-C levels. Furthermore, renal dysfunction in FLD frequently requires dialysis therapy in middle-aged patients and has a poor prognosis⁴⁾. Recently, immune-mediated acquired LCAT deficiency caused by LCAT autoantibodies was identified as a secondary LCAT deficiency. Although detailed reports are limited, it causes exacerbated conditions similar to FLD, such as low HDL-C levels and renal dysfunction⁵⁾.

Previous studies have shown that HDL has various functions including anti-atherosclerotic, anti-inflammatory, and antioxidant actions⁶⁾. Among these, the HDL cholesterol efflux capacity (CEC) is negatively associated with atherosclerotic cardiovascular disease events in both primary and secondary prevention settings^{7, 8)}. In general population studies, low HDL-C levels are frequently associated with a low HDL CEC⁷⁾. According to a recent clinical study, mutations in various genes that reduce the LCAT activity are strongly associated with atherosclerosis⁹⁾. Therefore, the HDL function may be crucial to the prognosis of immune-mediated acquired LCAT deficiency, in addition to LCAT activity and HDL-C concentrations.

In some cases of immune-mediated acquired LCAT deficiency, prednisolone therapy was shown to modulate LCAT antibodies, thereby improving the LCAT activity, HDL-C levels, and renal dysfunction^{5, 10)}. We previously reported two cases of acquired LCAT deficiency (HDL ＜5 mg/dL) with immune (immunoglobulin G4)-related LCAT autoantibodies¹¹⁾. One of them (Case 1 in the previous report) required prednisolone treatment due to a deterioration of medical conditions¹¹⁾ and was referenced against other cases with the same etiology^{5, 10)}. Because the initial therapeutic recovery process of the disease therapy was unclear, the current case report investigated the alteration of LCAT activity, lipid changes with their composition, HDL CEC, and renal dysfunction in the early stages of prednisolone treatment.

2.Methods

2.1. Blood and Urine Sampling and Biochemical Analyses

The patient provided his written informed consent for this case. Blood and urine samples were collected during inpatient and outpatient care and analyzed according to the clinical laboratory results. Blood and urine sampling, as well as biochemical analyses, were performed using the same method as in previous cases¹¹⁾. Blood samples were drawn during fasting (for several hours at admission and treatment-start day, or overnight on the other days) during the initial stage of treatment at 10 points (day of admission; 8 h after the start of treatment; and on days 2, 3, 6, 8, 10, 13, 22, and 29). As described previously¹²⁾, the serum LCAT activity was determined using an endogenous substrate as the cholesterol esterification rate. This method measured total LCAT activity, with a reference range of 82.1±23.1 nmol/ml/h.

Urine samples were collected before and after treatment at nine time points (day 20 before admission and days 0, 2, 3, 6, 8, 10, 13, and 29 of admission). A urine sample was collected on day 0 after hospitalization, overnight, and immediately before starting therapy. Urine protein levels were determined by a quantitative analysis (g/gCr).

2.2. HDL CEC

The HDL CEC at 10 sample points throughout the treatment course was measured as described previously^{7, 13)}. In brief, RAW 264 cells (RIKEN, Tsukuba, Japan) were radiolabeled with ³H-cholesterol (PerkinElmer, Waltham, MA, USA) for 24 h and then treated with 8-(4-chlorophenylthio)-cyclic AMP (cAMP) (Sigma, St Louis, MO, USA) for 18 h to activate adenosine triphosphate-binding cassette transporter A1 (ABCA1). Subsequently, 2.0% apolipoprotein B-depleted serum treated with the phosphotungstic acid-Mg precipitation method was added to the cells for 4 h, extracting cholesterol from the cells and into the supernatant. Finally, the efflux rates of ³H-cholesterol from cells to the medium were measured using a liquid scintillation counter. In the basic solution, serum-free medium with 0.2% fatty-acid-free bovine serum albumin (Calbiochem, Merck KGaA, Darmstadt, Germany) was used for radiolabeled treatment, stimulation, and serum addition in all HDL CEC experiments. The HDL CEC of each sample was measured in triplicate. The serum sample from a healthy subject served as the internal control in this study (reference range in quadruplicate, 10.22%±0.25%). HDL CEC is largely associated with the concentration of preβ HDL, which is affected by both the newly produced HDL by the ABCA1 pathway and lowered HDL catabolism by improving disturbed LCAT activity after treatment^{14, 15)}. Therefore, cAMP stimulation was adopted in the present case because it is important not to underestimate HDL cholesterol efflux mainly from the ABCA1 pathway and LCAT activity alteration by the treatment.

2.3. Lipoprotein Analyses

Serum samples from the present case and the control (a healthy subject) were fractionated using high-performance liquid chromatography with a gel filtration column (HPLC-GFC; LipoSEARCH; Immuno-Biological Laboratories Co., Ltd., Gunma, Japan)¹⁶⁾. Lipoproteins were divided into 20 subclasses: chylomicron (CM, fractions 1–2), very-low-density lipoprotein (VLDL, fractions 3–7: large, 3–5; medium, 6; small, 7), low-density lipoprotein (LDL, fractions 8–13: large, 8; medium, 9; small, 10–13), and HDL (fractions 14–20: large, 14–16; medium, 17; small, 18–20). HDL size was further divided into very large (fractions 14 and 15), large (fraction 16), medium (fraction 17), small (fraction 18), and very small (fractions 19 and 20). Simultaneously, these lipoprotein subclasses were analyzed using an enzymatic method to determine total cholesterol, triglycerides, phospholipids, and free cholesterol. Cholesteryl ester (CE) was calculated by subtracting the values of total cholesterol and free cholesterol.

3.Case Presentation

A 74-year-old Japanese man (Case 1 in the previous report¹¹⁾) experienced a sudden decrease in serum HDL-C level to 3 mg/dL 7 years before the present treatment, and this condition persisted. The cause was identified as “immune-mediated acquired LCAT deficiency” four years ago. Following the diagnosis, various fibrate treatments were initiated to reduce triglyceride levels, but HDL-C levels increased only slightly elevated, not enough to reach a normal HDL-C range (HDL-C fluctuation range 10–23 mg/dL), as reported in a previous case presentation¹¹⁾. For several years prior to treatment, the renal function deteriorated, with serum creatinine levels gradually fluctuating between 1.1 and 1.6 mg/dL, and urine protein was markedly aggravated ranging from 0.98–12.5 g/gCr. In comparison to previous cases^{5, 10)}, the current case began prednisolone treatment due to the continuation of exacerbated conditions, along with the prophylactic administration of sulfamethoxazole/trimethoprim. The patient was cautiously hospitalized for treatment in late 2021 because he was at increased risk due to underlying conditions and its relevant medications for coronary artery disease (telmisartan 10 mg/day, esomeprazole 20 mg/day, and cilostazol 100 mg/day), hypertension (benidipine 6 mg/day), diabetes mellitus (insulin and sitagliptin 50 mg/day), dyslipidemia (pemafibrate 0.2 mg/day), arrhythmia (apixaban 5 mg/day), hyperuricemia (benzbromarone 25 mg/day), and hypothyroidism (levothyroxine 25 µg/day). Table 1 displays the blood and urine profiles obtained during hospitalization. A low HDL-C level, LCAT activity below the measurement limit, and marked exacerbation of anemia and urine protein were detected for several years before treatment (Table 1).

Table 1.Blood and urine profile just before the treatment

(Reference range)				(Reference range)
T-Bil	0.9	mg/dL	(0.2-1.2)	ApoA-I	46	mg/dL	(119-155)
AST	33	U/L	(8-30)	ApoA-II	7.0	mg/dL	(25.9-35.7)
ALT	14	U/L	(5-35)	Fe	58	µg/dL	(80-160)
LDH	218	U/L	(100-225)	TIBC	311	µg/dL	(235-385)
ALP	67	U/L	(38-113)	Ferritin	266	ng/mL	(23-250)
γ-GTP	21	U/L	(7-70)	CRP	≦0.3	mg/dL	(≦0.3)
TP	6.9	g/dL	(6.5-8.2)	IgG	1323	mg/dL	(870-1700)
Alb	3.6	g/dL	(3.8-5.2)	LCAT activity	ND	nmol/ml/hr	(82.1±23.1)
BUN	23	mg/dL	(8.0-20)
Cre	1.88	mg/dL	(0.61-1.13)	WBC	3000	/µL	(3300-8600)
Na	141	mEq/L	(135-147)	RBC	352	×104/µL	(435-555)
K	4.2	mEq/L	(3.5-5.0)	Hb	8.9	g/dL	(13.7-16.8)
Cl	107	mEq/L	(98-108)	Hct	27	%	(40.0-50.0)
UA	7.0	mg/dL	(2.7-8.0)	MCV	76	fL	(83.6-98.2)
CK	124	U/L	(≦160)	MCH	25	pg	(27.5-33.2)
TC	156	mg/dL	(130-230)	MCHC	33	%	(31.7-35.3)
TG	428	mg/dL	(30-150)	Plt	19	×104/µL	(15.8-34.8)
HDL-C	18.7	mg/dL	(42-62)
Glu	151	mg/dL	(65-110)
HbA1c	4.3	%	(4.2-6.2)	U-protein＊	12.2	g/gCr	(＜0.15)

Abbreviations: T-Bil, total bilirubin; AST, L-aspartate: 2-oxoglutarate aminotransferase; ALT, L-alanine: 2-oxoglutarate aminotransferase; LDH, lactate dehydrogenase; ALP, alkaline phosphatase; γ-GTP, γ-glutamyl transferase; TP, total protein; Alb, albumin, BUN, blood urea nitrogen; Cre, creatinine; Na, sodium; K, potassium; Cl, chlorine; UA, uric acid; CK, creatine kinase; TC, total cholesterol; TG, triglyceride; HDL-C, high density lipoprotein-cholesterol; Glu, Glucose; HbA1c, hemoglobin A1c; Apo, apolipoprotein; Fe, ferrum; TIBC, total iron binding capacity; CRP, c-reactive protein; Ig, immunoglobulin; LCAT, lecithin–cholesterol acyltransferase; WBC, white blood cell count; RBC, red blood cell count; Hb, hemoglobin; Hct, hematocrit; MCV, mean corpuscular volume; MCH, mean corpuscular hemoglobin; MCHC, mean corpuscular hemoglobin concentration; Plt, platelet count; U-protein, urine protein, ND, not detected.

^＊U-protein was measured by quantitative analysis in 20 days before hospitalization (g/gCr).

Prednisolone oral therapy (30 mg/day) was initiated to restore LCAT activity the day after hospitalization. Blood samples were drawn during the initial stage of treatment at 10 points (admission day; 8 h after the start of treatment; and days 2, 3, 6, 8, 10, 13, 22, and 29). The patient was discharged within two weeks with no significant side effects and transitioned to outpatient care without changing any other medications. In Fig.1A, the results showed that LCAT activity was not detected from pretreatment to day 3 but gradually increased to the lower limit zone of normal (at day 8–22) and up to the proper level at day 29 (reference range, 82.1±23.1 nmol/ml/h)¹²⁾. Prednisolone therapy, however, caused an initial decline in HDL-C concentration until day 6, after which it improved to the normal range (Fig.1B). The apolipoprotein A-I (ApoA-I)/HDL-C ratio did not change significantly from 2.46 (admission day) to 2.96 (day 29). HDL CEC initially decreased but gradually returned to healthy levels (reference range, 10.22%±0.25%) (Fig.1C). Interestingly, both HDL CEC and HDL-C showed similar changes, consisting of an “initial dip and subsequent elevation,” with the former changing faster than the latter and reaching a plateau on day 22 while the latter was still in the upsloping phase (Fig.1D).

Fig.1. The one-month treatment progress by prednisolone therapy in the immune-mediated acquired LCAT deficiency case

(A) LCAT activity, (B) HDL-C concentration, (C) HDL CEC, (D) The combined data with HDL-C concentration and HDL CEC. LCAT, lecithin–cholesterol acyl transferase; HDL-C, high-density lipoprotein cholesterol; HDL CEC, high-density lipoprotein cholesterol efflux capacity. The reference range of HDL CEC was 10.22%±0.25% in this study. ^＊LCAT activity was very low because four points from days 0 to 3 were not detected.

A lipoprotein analysis was performed using high-performance liquid chromatography with a gel filtration column (HPLC-GFC)¹⁶⁾ at five points: day of admission (pretreatment), day 2 (lowest HDL CEC), day 6 (lowest HDL-C concentration), day 13 (recovery in progress), and day 29 (normal-range HDL-C concentration). Whole fractions of total cholesterol (TC), triglyceride, phospholipid, and free cholesterol at each time point and the control are displayed in Table 2 and Fig.2. On the day of admission and day 2, no CE was detected (Table 2, Fig.2A, and Fig.2B), which was consistent with the absence of LCAT activity (Fig.1A). FLD refers to specific lipoprotein characteristics, including (1) larger LDL size with different compositions, (2) increased phospholipid levels in particle sizes of small LDL (fraction 12)–large HDL (fraction 16), and (3) significantly decreased compositions of HDL fraction ¹⁶⁾. The current case demonstrated the same finding for the above characteristics: LDL with high triglyceride and phospholipid levels (fraction 8), increased phospholipid levels in fractions 12–16, and low component of HDL fraction analyzed by HPLC-GFC at pretreatment and day 2 (Fig.2A and 2B). After the treatment started, the original large LDL rich in triglycerides and phospholipids, a characteristic of FLD, changed to ordinary LDL rich in cholesterol, suggesting the recovery of β-LCAT activity (Fig.2A–2E). The HPLC-GFC analysis also provided detailed information about the recovery of the HDL composition (Fig.3). There was no marked difference in the very low values of TC and free cholesterol in the HDL fraction before treatment and on day 2 (Fig.3A and 3 B), indicating no LCAT activity. As shown in Table 2, the CE in the TC ratio (CE/TC ratio) increased in all fractions as LCAT activity increased. Phospholipid and CE levels in the HDL fraction increased significantly, suggesting an increase in α-LCAT activity (Fig.3C–3E) similar to the control level (Fig.3F). In terms of the HDL function, prednisolone therapy initially reduced the HDL CEC on day 2 before it returned to the pretreatment level on day 6, followed by improvement to levels comparable to those in healthy subjects (Fig.1C), thus allowing HDL to mature (Table 2 and Fig.3C–3E). Although there was no marked difference in the HDL composition measured by HPLC-GFC between pretreatment and day 2, phospholipids in the HDL fraction improved slightly by day 6. Therefore, these four components in the HDL fraction analyzed by HPLC-GFC could not fully explain the improvement in HDL CEC.

Table 2.Fluctuations of lipid-constituted factors by HPLC-GFC analysis at 5 featured-points during the one-month treatment

		Admission day	Day 2	Day 6	Day 13	Day 29	Control
Total cholesterol
All fractions	mg/dL	89.6	88.9	127.8	216.5	308.6	131.3
CM (Fraction 1 – 2)	mg/dL	13.0	15.9	13.4	8.4	3.2	0
VLDL (Fraction 3 – 7)	mg/dL	39.5	40.3	49.4	61.6	86.1	8.6
LDL (Fraction 8 – 13)	mg/dL	29.7	25.9	58.3	129.8	185.5	67.0
HDL (Fraction 14 – 20)	mg/dL	7.4	6.7	6.7	16.7	33.8	55.6
Triglyceride
All fractions	mg/dL	275.8	247.1	299.8	259.3	225.3	42.9
CM (Fraction 1 – 2)	mg/dL	32.5	36.7	34.9	27.5	8.3	0.2
VLDL (Fraction 3 – 7)	mg/dL	151.1	137.4	147.8	140.8	123.1	9.9
LDL (Fraction 8 – 13)	mg/dL	87.1	68.3	106.5	78.8	79.7	18.8
HDL (Fraction 14 – 20)	mg/dL	5.1	4.7	10.5	12.2	14.3	14.0
Phospholipid
All fractions	mg/dL	267.2	248.6	238.6	269.2	363.9	184.3
CM (Fraction 1 – 2)	mg/dL	20.0	22.6	18.8	9.5	3.9	0.1
VLDL (Fraction 3 – 7)	mg/dL	90.9	89.4	77.7	71.0	83.1	5.8
LDL (Fraction 8 – 13)	mg/dL	98.6	82.3	92.2	120.3	158.0	55.5
HDL (Fraction 14 – 20)	mg/dL	57.6	54.2	49.9	68.4	118.8	122.9
Free cholesterol
All fractions	mg/dL	89.3	79.0	70.5	76.5	98.9	29.6
CM (Fraction 1 – 2)	mg/dL	11.1	12.3	9.4	4.5	1.4	0
VLDL (Fraction 3 – 7)	mg/dL	38.0	35.8	30.0	27.2	31.5	1.9
LDL (Fraction 8 – 13)	mg/dL	31.6	24.2	28.2	41.4	60.1	21.1
HDL (Fraction 14 – 20)	mg/dL	8.7	6.7	2.8	3.3	5.9	6.7
CE/TC
All fractions	%	0.3	11.1	44.8	64.7	68.0	77.4
CM (Fraction 1 – 2)	%	14.0	23.0	29.4	46.7	55.9	23.0
VLDL (Fraction 3 – 7)	%	4.0	11.2	39.3	55.8	63.5	78.4
LDL (Fraction 8 – 13)	%	0	6.5	51.6	68.1	67.6	68.6
HDL (Fraction 14 – 20)	%	0	0.2	57.8	80.0	82.5	88.0

Abbreviations: HPLC, high-performance liquid chromatography; CM, chylomicron; VLDL, very-low-density lipoprotein; LDL, low-density lipoprotein; HDL, high-density lipoprotein; CE/TC ratio, cholesteryl ester/total cholesterol ratio.

Fig.2. An HPLC-GFC lipoprotein analysis at five featured-points during the one-month prednisolone treatment

(A) Admission day at pretreatment, (B) Day 2 at the lowest HDL CEC, (C) Day 6 at the lowest HDL-C concentration, (D) Day 13 at the recovery in progress, (E) Day 29 at the HDL-C concentration with normal range, (F) the control. HPLC-GFC, high-performance liquid chromatography with gel filtration column; CM, chylomicron; VLDL, very-low-density lipoprotein; LDL, low-density lipoprotein; HDL, high-density lipoprotein.

Fig.3. HDL-specific fractions in the HPLC-GFC analysis at five featured-points during the one-month treatment

(A) Admission day at pretreatment, (B) Day 2 at the lowest HDL CEC, (C) Day 6 at the lowest HDL-C concentration, (D) Day 13 at the recovery in progress, (E) Day 29 at the HDL-C concentration with normal range, (F) the control. HPLC-GFC, high-performance liquid chromatography with gel filtration column; HDL, high-density lipoprotein. HDL size indicates very large (fraction 14 and 15), large (fraction 16), medium (fraction 17), small (fraction 18), and very small (fraction 19 and 20).

Urine protein levels increased significantly to 12.2 g/gCre prior to treatment (Table 1). However, proteinuria spontaneously improved with hospitalization alone (the point with an asterisk in Fig.4) and was further improved by treatment during hospitalization (range 1.3–2.6 g/gCre). However, the urine protein level again increased at day 29 (5.4 g/gCr), although still remaining better than the pre-hospitalization level (Fig.4). Serum creatine levels remained unchanged before and after the one-month steroid treatment (range 1.55–1.95 mg/dL).

Fig.4. Urine protein changes before and after the one-month steroid treatment

Urine protein was calculated by a quantitative analysis (g/gCr). ^＊Urine protein on day 0 was measured after hospitalization overnight and immediately before therapy.

4.Discussion

The present patient with immune-mediated acquired LCAT deficiency received prednisolone therapy because of worsening disease conditions. The initial one-month treatment increased LCAT activity and improved the lipid profile from FLD-like to the normal range. The recovery of HDL from low to normal levels was also demonstrated by conventional and HPLC analyses. The HDL CEC improved to normal as the HDL-C concentration increased. However, the improvement in the HDL CEC was found to be more sensitive and faster than that of HDL-C. Therefore, normal HDL-C levels may not be a crucial treatment target in patients with immune-mediated acquired LCAT deficiency who require treatment. Urine protein was significantly improved by the treatment, although serum creatine levels remained unchanged. To our knowledge, this is the first case report in which prednisolone therapy restored the HDL function during short-term treatment.

Immune-mediated acquired LCAT deficiency may have two components: LCAT deficiency and autoimmune-like conditions that produce LCAT autoantibodies. As a result, the course after the disease onset is expected to vary, and some cases range from spontaneous improvement¹⁷⁾ to a strong need for prednisolone therapy^{5, 10)}. In the current case, LCAT activity was restored to normal levels approximately 10 days after the start of steroid therapy (Fig.1A), which was nearly identical to the duration (2 weeks) reported in a previous paper¹⁰⁾. Furthermore, the LCAT activity detected by the endogenous substrates method almost completely recovered, thus suggesting that both the α- and β-LCAT activities had improved (Table 2 and Fig.2). In this case, the LCAT activity and HDL-C concentration increased to normal levels after one month of treatment. However, the duration of HDL recovery may vary owing to differences in these two aspects in other cases.

Treatments for FLD have been studied and developed to improve LCAT protein and activity, HDL-C levels, and prevent renal dysfunction¹⁸⁾. Recent human treatment reports for FLD include recombinant human LCAT (rhLCAT) infusions¹⁹⁾ and gene therapy with autologous implantation of adipocytes genetically expressing human LCAT²⁰⁾. A previous study using rhLCAT demonstrated remarkable rescue of LCAT activity and HDL-C elevation in FLD cases. In the same study, HDL-C concentration significantly surged from 0.5 h to a maximum of 6 h immediately after the start of rhLCAT infusion in a dose-dependent manner¹⁹⁾. However, the current case required six to eight days to recover LCAT activity and approximately a month to increase HDL-C levels (Fig.1). The presence of LCAT autoantibodies was demonstrated by qualitative testing before and after HDL recovery in the current case¹¹⁾ and the recently reported case¹⁷⁾ of immune-mediated acquired LCAT deficiency. Similarly, the recent case found almost the same volume of LCAT autoantibodies in the blood both before and after HDL-C recovery. Based on these findings, the inhibition of LCAT activity may indicate that the affinity of the autoantibody for LCAT needs to be considered in addition to its volume. Thus, it may take more time for prednisolone to increase HDL-C concentration by modulating LCAT autoantibodies, compared to FLD.

Cholesterol efflux was found to have different outcomes in two FLD cases with extremely low HDL-C levels (2–4 mg/dL) compared to normal subjects^{21, 22)}. In contrast, rhLCAT administration resulted in HDL-C elevation after 1 h, and the HDL CEC increased after 12 h in patients with low HDL-C levels and coronary heart disease²³⁾. In the present case, the HDL CEC was low, with undetectable LCAT activity and a low HDL-C concentration of 18.7 mg/dL prior to therapy (Fig.1). Prednisolone treatment significantly increased HDL CEC to a healthy level after approximately three weeks, and interestingly, this recovery occurred after LCAT activity recovery but before HDL-C concentration recovery (Fig.1). Overall, the recovery of LCAT activity may affect the HDL CEC in patients with immune-mediated acquired LCAT deficiencies. In circulating HDL particles, there were no consistent changes between HDL CEC and HDL composition at the three time points (pretreatment, day 2, and day 6), as determined by an HPLC-GFC analysis with TC, triglyceride, phospholipid, and free cholesterol. Consequently, other HDL-associated proteins may also be involved in HDL CEC recovery. In this disease, HDL CEC appears to be lower when LCAT activity and HDL levels are significantly reduced by LCAT autoantibodies. However, prednisolone-based treatment has been reported to be effective for exacerbated conditions of immune-mediated acquired LCAT deficiency^{5, 10, 24)}. Interestingly, the present case showed that HDL CEC may be improved to the normal range even if HDL-C concentrations do not return to normal levels, such as above 30 mg/dL (Fig.1). In HDL recovery, the time course may be as follows: (1) premature HDL (preβ HDL) produced by the ABCA1 pathway is metabolized and excreted before the therapy (little LCAT activity); (2) after starting the treatment, preβ HDL increases following LCAT activity improvement due to modulation of LCAT autoantibodies; and (3) mature HDL was gradually constructed from preβ HDL due to sufficient LCAT activity. As a result, (4) HDL-C levels increased. From this process, the improvement of HDL CEC, whose function is known to be closely related to the preβ HDL concentration¹⁴⁾, may be faster than that of HDL-C. Conversely, glucocorticoids have been reported to downregulate cholesterol efflux and ABCA1 expression in macrophages for a short duration in vitro²⁵⁾. The same study further showed that the interaction between glucocorticoids and glucocorticoid receptors might play a role in reducing the HDL CEC through the ABCA1 pathway using a gene manipulation experiment²⁵⁾. As such, the initial decrease in the HDL CEC and HDL-C levels may have been affected by prednisolone treatment in the present case.

Treatments for renal failure caused by low LCAT activity in FLD include low-fat diet, dialysis, kidney transplantation, and gene therapy³⁾. In particular, renal failure with proteinuria in FLD is thought to be associated with low LCAT activity⁴⁾. Renal dysfunction can relapse in FLD with low LCAT activity, even after kidney transplantation³⁾. In contrast, a case report suggested that glucocorticoid treatment improved proteinuria in FLD²⁶⁾. According to a case report, the effect was most likely caused by inhibiting inflammation in the kidney rather than by improving LCAT activity. In the case of immune-mediated acquired LCAT deficiency, a renal biopsy revealed a significant reduction in foam cells in the kidney following prednisolone treatment¹⁰⁾. In the current case, prednisolone treatment significantly improved urine protein levels (Fig.4), which most likely resulted in an improvement in both LCAT activity and kidney inflammation. However, the therapy in this case did not completely correct the proteinuria after one month of treatment. A recent study revealed that the association between HDL CEC and incident cardiovascular events is recognized in subjects with mild CKD, although this association disappears among patients with advanced CKD²⁷⁾. From this perspective, the recovery of renal dysfunction by treatment may improve HDL CEC and increase the importance of HDL CEC in this case.

The present patient’s hypothyroidism was not that of typical Hashimoto’s disease since the patient was negative for diagnostic autoantibodies of Hashimoto’s disease, such as anti-thyroid peroxidase antibody or anti-thyroglobulin antibody. Therefore, the association between autoimmune reactions to LCAT and Hashimoto’s disease remains unclear.

Several limitations associated with the present study warrant mention. First, we should bear in mind that some patients with immune-mediated acquired LCAT deficiency showed spontaneous improvement. Second, since the patient in this case was an elderly man with multiple medical complications, including coronary heart disease and diabetes mellitus, and receiving numerous medications, our findings should be interpreted cautiously when being applied to those who differ in sex, age, or complications. Third, in this case, LpX cannot be revealed because the HPLC-GFC method cannot detect any abnormal lipoproteins which are consistent with the LpX definition¹⁶⁾.

In conclusion, one-month prednisolone therapy restored three HDL-related parameters: LCAT activity, HDL CEC, and HDL-C concentration. Prednisolone therapy significantly improved proteinuria but failed to achieve normal levels, suggesting that nomalization of the kidney function will take longer to achieve and require more than simply HDL restoration.

Acknowledgements

This work was supported in part by funding from Fukuoka University (Grant No. 207108) (T.K.). This research was partially supported by AMED under Grant Number JP22ek0109461(K.Y.). This work was partially supported by a Health, Labour, and Welfare Science Research Grant for Research on Rare and Intractable Diseases (H30-nanji-ippan-003)(K.Y.). Serum LCAT activity was determined using CellGenTech, Inc. (Chiba, Japan).

Conflict of Interest

Y.U. contracted with RAYDEL Australia Pty. Ltd. as a research consultant and received research grants from Asahi Kasei Co. LTD., Mizuno Co. LTD., Scuba diving・Air. LTD., and RAYDEL Korea Co. Ltd. This work was partially supported by joint research funding from CellGenTech, Inc. (K.Y.), and a contract research grant from CellGenTech, Inc. (M.K.). The authors declare that they have no conflicts of interest.

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