Surgery

Individual biomarkers in the blood are not yet applicable in diagnosing complicated appendicitis: A scoping review

a b s t r a c t

Background: Appendicitis is one of the most common surgical emergencies globally and it can both be difficult to diagnose but also to differentiate complicated from uncomplicated appendicitis preoperatively. The objective of this scoping review was to develop an overview of biomarkers in blood discriminating complicated from uncom- plicated appendicitis and characterize their applicability in an acute setting including time, cost, and analysis technique required as well as their individual precision.

Method: This scoping review was reported in accordance with PRISMA-ScR. The included studies had to report on biomarkers measured in the blood for at least ten patients with suspected appendicitis. A systematic literature search was conducted on August 28, 2022, in PubMed and Embase but restricted to articles published in January 2000 and onwards. A protocol was uploaded to Open Science Framework prior to data extraction.

Results: A total of 65 biomarkers were included from 52 studies, covering 14,312 patients. There was 60% routine- and 40% novel biomarkers based on the reported analysis technique. The most frequently investigated biomarkers within each group were white blood cell count and procalcitonin. The routine biomarkers were of low financial cost but poor diagnostic accuracy with sensitivity ranging between 15 and 100% and specificity between 27 and 100%. Novel markers were costly ranging from 275 to 800$, and their diagnostic accuracy was based on limited population sizes (median 34 patients) and reported for only 5% of the novel markers.

Conclusion: Routine biomarkers were applicable in an acute setting but had poor diagnostic accuracy. Novel bio- markers are being investigated for potential, but the concept is still premature due to lack of diagnostic accuracy studies reporting cost-benefit for individual markers and whether they can be applied in an acute setting.

(C) 2023

  1. Introduction

Acute appendicitis is recognized as the most common cause of emer- gency abdominal surgery and with an annually rising incidence in newly industrialized countries in Asia, the Middle East, and Southern America [1]. The severity of appendicitis can be divided into two distinct groups, uncomplicated and complicated appendicitis [2]. The latter is defined intraoperatively as a gangrenous or perforated appendix or an abscess formation [3]. The diagnosis of appendicitis remains challenging and relies on clinical symptoms [4]. These are of poor predictive value when applied as a singular diagnostic tool, but supplementary inflam- matory biomarkers including the complete blood count with differential count and imaging modalities, where accessible, can help support the diagnosis. However, CT scans express low accuracy in discriminating be- tween complicated and uncomplicated appendicitis [5] and is costly. In comparison to diagnostic imaging, biomarkers are more easily

* Corresponding author.

E-mail address: [email protected] (B. Sikander).

accessible at almost every hospital and provide a less invasive measure- ment for a systemic response caused by the inflammation of the appen- dix. Ideally, biomarkers function as a precursor for a pathological condition that is difficult to observe [6] and could therefore be useful to predict the severity of appendicitis. Treatment of complicated appen- dicitis relies solely on the removal of the inflamed appendix by either open or laparoscopic appendectomy. However, the timing of the surgi- cal intervention is an important factor to avoid postoperative complica- tions, hence, the reason why a Timely diagnosis is crucial [7].

This scoping review aimed to develop an overview of biomarkers in the blood to discriminate between complicated and uncomplicated appendicitis preoperatively. Furthermore, we aimed to assess their applicability in an acute setting by characterizing the time, cost, and analysis technique required as well as the precision of each biomarker.

  1. Methods

We reported this scoping review according to the Preferred Reporting Items for Systematic reviews and Meta-Analysis extensions

https://doi.org/10.1016/j.ajem.2023.02.016

0735-6757/(C) 2023

for Scoping Reviews (PRISMA-ScR) [8] and Joanna Briggs Institute’s manual for reporting scoping reviews [9]. A protocol was registered and uploaded to the Open Science Framework [10]. The focus was on biomarkers that differentiated complicated from uncomplicated appen- dicitis and were measured in blood samples (blood, serum, or plasma). Studies concerning participants with suspected appendicitis and subse- quently histopathological verification of an inflamed or normal appen- dix were considered for inclusion. The patients with complicated appendicitis had to be assessed at surgery based on the intraoperative appearance and defined as at least gangrenous and perforated appen- dix, but also abscess or other findings of complicated disease if men- tioned [3]. Patients with a complicated appendicitis diagnosis based on other reports than the intraoperative description were not included in this study, e.g. imaging, cultures, or histopathological reports. We in- cluded cohort studies, both retrospective and prospective, consisting of a minimum of ten participants with suspected appendicitis. Only full- text original articles published in English, German, French, Urdu, and

the Scandinavian languages were eligible for inclusion. Studies pub- lished from the year 2000 and onwards were considered for inclusion to maintain contemporary clinical management. There was no restric- tion in terms of age, sex, ethnicity, or race of patients. Articles in lan- guages other than the above-mentioned, published before the year 2000, and consisting of less than 10 participants with suspected appen- dicitis were excluded.

The pilot searches included terms related to either appendicitis or appendectomy and terms for biomarker, predictor, or blood. The final broad systematic literature search was conducted with the assistance of a research librarian [11] who was consulted regarding both develop- ments of the search string as well as the choice of the databases. The last date of the search was August 28, 2022, in two databases PubMed and Embase. For each database, the search string was adapted. For PubMed, the following search string was used: (Appendicitis OR appendicitis [MeSH Terms] OR appendectomy OR appendicectomy OR appendec- tomy [MeSH Terms]) AND (biomarker OR biomarker [MeSH Terms]

Image of Fig. 1

Fig. 1. Flow diagram of the study Selection process. n: number.

OR marker OR “blood” OR blood [MeSH Terms] OR serum OR plasma OR predictor). The modified search string for Embase, can be found in the protocol [6]. Following the search in both databases, the records were uploaded to Mendeley (version 1.19.8, Elsevier, UK) to check for and re- move duplicates, subsequently exporting them for screening to the on- line screening tool Covidence [12]. The screening process consisted of three steps before deciding which studies to be included. Firstly, two au- thors independently screened titles and/or abstracts for all records pro- duced by the search string. Secondly, the eligible reports were screened in full text by the same two authors independently. Lastly, the first au- thor conducted a snowball search to explore the reference lists of the in- cluded studies to identify other studies eligible for inclusion [11]. This was supplemented with a screening of the reference lists of identified systematic reviews.

Prior to the charting of data, a pilot test was conducted on ten articles independently by two authors. Any conflicts were resolved internally within the author group by discussion. The relevant variables were ex- tracted to an excel sheet twice to ensure uniform extraction, and dis- agreements were resolved within the author group by discussion. All the extracted variables can be seen in the protocol [6]. The extracted data items included details on the study characteristics such as the study design, country of origin, and language. For patient characteristics, the distribution of age amongst the included population, the distribu- tion of sex amongst the complicated appendicitis population, and the definition of complicated appendicitis for each of the included studies were extracted. Information regarding storage, analysis technique, time taken before results were received, and the cost was extracted for all the biomarkers. The accuracy including the negative predictive value, positive predictive value, sensitivity, and specificity when com- pared between complicated and uncomplicated appendicitis was also extracted. Biomarkers were categorized as either routine or novel de- pending on the analysis methods, e.g. routine if the analysis method for a biomarker was described as “routine laboratory results” in at least one of the studies reporting on that specific biomarker. The bio- markers were further divided into groups according to the specific area of functional measure, e.g. inflammatory response, coagulation profile, kidney- and liver function test, bacterial or viral tests, and others.

  1. Results

The selection of the studies is presented in Fig. 1. The search was conducted across two different databases yielding 9689 records. After removing duplicates, a total of 7790 records went on to the title and ab- stract screening. Finally, a total of 52 original articles, reporting on 65 different biomarkers were included in this scoping review [13-64].

    1. Characteristics of studies and biomarkers

The summary study characteristics can be seen in Table 1. The study designs were 52% prospective cohort studies [13,14,17,20,22,24- 26,28,30,31,33,36-39,41,43,46,47,49-53,60,64] and 48% retrospective

cohort studies [15,16,18,19,21,23,27,29,32,34,35,40,42,44,45,48,54-

59,61-63]. The studies were conducted in 23 countries mainly in Turkiye 33%, Pakistan 8%, and Sweden 6% with a total population of 14,312 unique patients with suspected appendicitis included across studies as two reports had overlap in populations [18,19]. In terms of age, the majority of the studies included primarily adult patients 67%, followed by pediatric patients 23%, and in 4% of studies, patients of all ages were included. In 6% of studies, the age group was not specified. Complicated appendicitis was operatively diagnosed in 4005 patients (28% of the total population). The size of the complicated appendicitis population within each included study ranged from 7 [27] to 443 [15] patients. Sex was reported in 71% of studies, reporting a male domi- nance of 61% for patients with complicated appendicitis. The definitions

Table 1 Summary characteristics of the 52 included studies reporting on biomarkers in the blood presented as numbers (n) and percentages of included studies.

A: Necrosis, free pus, and peritonitis, B: Representing different types of funding

including institutional (n = 4), industrial (n = 2), and unspecified (n = 2). Characteristic n %

Study design

Retrospective cohort study 25 48

Prospective cohort study 27 52

Country

Turkiye 17 33

Pakistan 4 8

Sweden 3 6

Korea 3 6

Spain 3 6

Other 22 42

Age

Adult patients 35 67

Pediatric patients 12 23

All ages 2 4

Not specified 3 6

Definition of complicated appendicitis

Gangrene and perforated appendix 24 46

Gangrene, perforated appendix, and abscess 26 50

Gangrene, perforated appendix, and otherA 2 4

Any conflict of interest 0 0

Received funding 8B 15

of complicated appendicitis in the included studies are shown in Table 1.

A total of 65 biomarkers, shown in Fig. 2, were divided depending on the analysis technique into 60% routine and 40% novel biomarkers. Ala- nine aminotransferase and aspartate aminotransferase were assumed as routine biomarkers, though it was not directly reported. The bio- markers were further grouped based on what functional measure they tested. Markers regarding the inflammatory response made up the larg- est proportion 49%, followed by coagulation profile markers at 23%, liver function test at 15%, kidney function test at 8%, bacterial or viral markers at 5%, and the remaining 8% marked as others. The most frequently re- ported routine biomarkers were C-reactive protein and white blood cell count, which was investigated in a total of 30 studies covering 2093 pa- tients with complicated appendicitis [13,14,17,19,20,22-27,30,32- 34,36,37,41,42,49-52,54,55,58,60,61,63,64] and 34 studies covering 2761

patients with complicated appendicitis [13,14,16,17,19,21,22,24,26,27,32- 37,39,41,42,44,46,50-53,55,57-64], respectively.

    1. Applicability of the biomarkers in an acute setting

The applicability of identified biomarkers in an acute setting was de- termined based on their handling and analysis technique along with the time to results and reported cost of implementing these individually. For the routine biomarkers, the analysis technique was reported in 62% of biomarkers, but only for 11% of biomarkers the time needed for the handling, processing, and receiving of results was reported [37- 39,44,45,50,63]. The median time for results was one hour, ranging from 1 to 24 h. The cost was not reported for any of the routine biomarkers.

For the novel biomarkers, handling and analysis technique were re- ported for all, see Table 2. The most frequently applied analysis tech- nique was simplex enzyme-linked immunosorbent assay (ELISA) (33%), whereas multiplex ELISA was applied in 16% of biomarkers, and the remaining markers used various methods (Table 2). Even though the technique for analysis was reported for all novel biomarkers, the time to results was not reported for 81% of these, but a reference time for each analysis technique could be extrapolated from an external source via the reported product details [65]. The reported time for re- sults for the novel biomarkers was 4 h. Similar to the routine biomark- ers, the cost was not reported directly in the included studies but based on the referral details for the analysis methods [65] the cost of

Image of Fig. 2

Fig. 2. An illustration of routine (green) and novel (red) biomarkers divided into respective types of tests (dotted circles). The circle areas illustrate the relative distribution of biomarkers in percent.

*Alanine aminotransferase/Aspartate aminotransferase were not reported as routine in the study but are assumed as such. CD: cluster of differentiation, MMP: matrix metalloproteinase, TNF-?: tumor necrosis factor-?, IL: interleukin.

the performed tests could be extrapolated from an external source and ranged from $275 to $800, see Table 2. Storage requirements were poorly reported in all 52 studies and could therefore not be compared between the routine and novel biomarkers.

    1. Accuracy

We investigated the individual diagnostic accuracy in discriminating complicated from uncomplicated appendicitis. For 73% of the routine biomarkers, one or more measures of accuracy were reported. Only 26% (n = 29) of these had an accuracy measured for complicated versus uncomplicated appendicitis (Table 3). The reported accuracy for the routine markers for thrombocyte to neutrophil ratio and total mast cells depicted these as superior diagnostic predictors, but only one study reported on each of these markers and their complicated popula- tion comprised of 18 patients [36]. The accuracy of routine biomarkers such as Neutrophil to lymphocyte ratio and C-reactive protein were in- vestigated and reported in five and four different studies, respectively, and presented with mediocre precision values. C-reactive protein had

a median [range] sensitivity of 71% [42-97%] and specificity of 80% [54-90%], whereas neutrophil to lymphocyte ratio had a median sensi- tivity of 71% [61-78%] and a specificity of 67% [49-77%] spread over a larger number of patients with complicated appendicitis. For the novel biomarkers, an accuracy measure was reported in 67% of biomarkers, but merely 5% of these had accuracy reported for complicated versus uncomplicated appendicitis. For the novel markers, only procalcitonin and cancer antigen 125 presented with good precision measures, but similarly to the previously mentioned routine markers, each of these was only reported in one study each with a population group with com- plicated appendicitis of 42 and 10 participants. Hence, the overall accu- racy for these markers was imprecise.

  1. Discussion

This scoping review provides an overview of the 65 biomarkers in blood that have been evaluated preoperatively to diagnose compli- cated appendicitis. More than half of the biomarkers were routine laboratory tests and can therefore be assumed to be applicable in

Table 2 The characteristics of analysis techniques for the novel biomarkers in the blood presented along with the reported time for results and referenced time found through external links. ELISA: enzyme-linked immunoassay, -: not reported.

Novel biomarkers

Analysis technique

Time in hh:mm

Cost, $

Reported

[69]

[69]

activated partial thromboplastin time

Manual

HemosIL Kit

04:00

Alkaline phosphates

Simple

ELISA

04:45

630

Calprotectin

Simple

ELISA

04:45

600

Cancer antigen 125

Simple

ELISA

04:45

800

Cluster of differentiation 44

Manual

Immunological dot-blot assay

Cluster of differentiation 64

Simple

ELISA

04:00

04:45

800

D-dimer

Simple

ELISA

04:45

600

Endotoxin

Manual

Limulus amebocyte lysate

00:15

80

Fibrinogen

Manual

HemosIL Kit

04:00

Immunoglobulin E

Simple

ELISA

04:45

600

Interleukin-1?

Simple

ELISA

04:45

605

Interleukin-4

Multiplex

ELISA

01:30

275

Interleukin-6

Multiplex

ELISA

01:30

305

Interleukin-8

Simple

ELISA

04:45

600

Interleukin-9

Multiplex

ELISA

01:30

275

Interleukin-13

Multiplex

ELISA

01:30

275

Matrix metalloproteinase 1-13

Multiplex

Magnetic bead-based ELISA

01:30

275

matrix metalloproteinase 9

Multiplex

Magnetic bead-based ELISA

01:30

275

Neopterin

Manual

DRG Neopterin 96-test

02:30

800

Plasminogen activator inhibitor type 1

Multiplex

Magnetic bead-based ELISA

01:30

275

Procalcitonin

Manual

LUMItest PCT assay

01:15

800

Prothrombin

Manual

HemosIL Kit

04:00

Thrombin time

Manual

HemosIL Kit

04:00

Tissue inhibitors of metalloproteinases 1-4

Multiplex

Magnetic bead-based ELISA

01:30

275

Tissue inhibitors of metalloproteinase 1

Multiplex

Magnetic bead-based ELISA

01:30

275

Tumor necrosis factor ?

Simple

ELISA

04:45

580

the acute setting, although it was not reported directly. Novel bio- markers lacked direct reporting on the analysis technique but pro- vided enough details to retrieve the information from an external source. However, the analysis techniques for novel biomarkers were costly and time-consuming, thus, not applicable in an acute setting. Reporting on accuracy for both routine and novel biomark- ers in discriminating complicated from uncomplicated appendicitis lacked in many studies. The novel biomarkers were investigated in single studies and within small populations, while the routine bio- markers were investigated in more studies and with larger popula- tions of complicated appendicitis, but either way the individual accuracy for these biomarkers was poor.

The strengths of this scoping review include the sound systematic search, the rigor methodology [7,9], and transparent reporting [8]. A scoping review approach was chosen due to the intention of investigat- ing previous literature on all diagnostic markers for complicated appen- dicitis and their relative applicability [65]. The previous literature helped provide insight into the lack of reporting in areas such as cost- benefit and diagnostic accuracy specifying cutoff values for each of these biomarkers in differentiating complicated from uncomplicated appendicitis [66]. However, some limitations were present. Due to the restrictions applied in our eligibility criteria in terms of Publication year and languages, there is a risk of language and selection bias. The definition of complicated appendicitis based on intraoperative findings

Table 3 Accuracy measures for routine and novel biomarkers in the blood are either presented as the specific accuracy measure or median [range] presented in percentages. n: number, PPV: pos- itive predictive value, NPV: negative predictive value, -: not reported.

Biomarkers with study references

Studies, n

Patients with complicated appendicitis, n

Sensitivity

Specificity

PPV, %

NPV, %

Routine biomarkers

Sodium [15,45]

2

[20-443]

53 [31-74]

74 [51-96]

Potassium [45]

1

20

78

27

Urea [45]

1

20

65

35

Creatinine [45]

1

20

65

30

mean platelet volume [62,63]

2

[47-208]

54 [34-73]

63 [57-69]

85

16

White blood cells [16,36,44,63]

4

[18-208]

73 [46-89]

73 [42-92]

65 [35-96]

50 [21-80]

Neutrophil [54,63]

2

[208-254]

58 [45-71]

67 [54-80]

92 [91-92]

36 [21-51]

C-reactive protein [30,54,58,63]

4

147 [42-254]

71 [42-97]

80 [54-90]

85 [72-89]

37 [18-96]

Neutrophil percentage [16,44]

2

[36-191]

80 [62-97]

43 [32-54]

79

34

Neutrophil to lymphocyte ratio [16,36,40,54,62]

5

[18-254]

71 [61-78]

67 [49-77]

51 [21-82]

69 [48-90]

Thrombocyte to neutrophil ratio [36]

1

18

94

94

Thrombocyte to lymphocyte ratio [36]

1

18

61

58

Total mast cells [36]

1

18

100

100

Mast cells degranulation [36]

1

18

100

100

Platelet to lymphocyte ratio [16]

1

36

42

86

Platelet distribution width [63]

1

208

15

85

83

16

Novel biomarkers Procalcitonin [30]

1

42

95

74

68

89

Prothrombin [44]

1

191

76

43

33

82

Fibrinogen [44]

1

191

63

75

49

85

Thrombin time [44]

1

191

50

69

38

79

Cancer antigen 125 [28]

1

10

60

100

100

was required, and articles with only histopathological verification were excluded. This led to exclusion of several reports, however, uniformed the reporting on complicated appendicitis in this review. The excluded articles mainly reported on routine biomarkers already included in this scoping review and very few studies on novel biomarkers that expressed good accuracy in discriminating complicated from uncompli- cated appendicitis such as fibrinogen [67] and thiol-disulfide [68] but did not meet the inclusion criteria. Cost and time to results were not re- ported in many articles, and an external source [69] had to be investi- gated to derive the values for the reported products.

Complicated and uncomplicated appendicitis are different disease entities with similar symptoms, but different treatment options [70- 72]. Early identification of complicated appendicitis is crucial to prevent in-hospital delay as it can lead to postoperative complications such as sepsis, Wound infections, fistulas, and reintervention [7]. The current ev- idence suggests that diagnostic imaging is efficient in diagnosing appen- dicitis [5,73] but that it lacks precision in differentiating complicated from uncomplicated appendicitis. Also, diagnostic imaging is a relatively expensive modality and is not practically applicable in all areas of the world. On the contrary, routine biomarkers including white blood cell count, C-reactive protein, and Neutrophil count are associated with low financial cost and efficient time to results, but the diagnostic accu- racy individually remains poor [74]. Novel biomarkers are emerging, but they are not common in clinical practice for the diagnosis of compli- cated appendicitis mainly due to limited evidence regarding their diag- nostic accuracy, significant costs, and the requirement of specialized personnel for the handling of the analysis technique. The median cost for an ELISA simplex test was 600$ and it took close to 5 h before the re- sults were final, however, ELISA multiplex was relatively cheaper and less time-consuming [69]. Only two of the 15 studies testing for novel biomarkers opted for the ELISA multiplex technique, both studies were relatively new and focused on a pediatric population only, and nei- ther reported accuracy measures [31,46]. Finally, there is a deficit in re- search exploring the applicability of these novel biomarkers in acute settings for the diagnosis of complicated appendicitis.

Clinicians’ early and accurate diagnosis of complicated appendicitis is of utmost importance to ensure that patients receive the most appro- priate and effective treatment, minimizing the risk of postoperative complications and promoting a successful recovery. The results of our study indicate that no single biomarker can be used by clinicians to re- liably differentiate between patients with complicated and uncompli- cated appendicitis. Biomarkers in the blood could instead be used by clinicians in the preoperative diagnosis of complicated appendicitis when combined with other variables [75]. Either as a panel consisting of more than one biomarker or together with other information used in the diagnostic process such as symptoms, physical examination, and Imaging techniques such as ultrasound or CT scans as done in diag- nostic tools or scoring systems [76,77]. It is also important to assess the willingness of clinicians to adopt these biomarkers and diagnostic tools in comparison to their custom Diagnostic approach. This is because the translation from research to clinical practice can be challenging. It is es- sential to understand the barriers to implementation to ensure the ef- fective adoption of these new diagnostic methods in the clinical setting [78].

In conclusion, this scoping review provides an overview of the diag- nostic accuracy of routine and novel biomarkers in the blood, including their cost and individual requirements, and it, therefore, assists the tran- sition of research on appendicitis from the laboratory to clinical settings. Routine biomarkers were accessible and inexpensive but lacked accu- racy. Conversely, novel biomarkers were unfeasible in an acute setting and expensive but may have better accuracy. The design of and reporting on future studies on biomarkers for the diagnosis of compli- cated appendicitis must prioritize simplicity and practicality in terms of time, ease of handling, and cost-effectiveness. This scoping review has consolidated the results of various research efforts and could thereby lead to a more relevant and effective utilization of time and

resources for future studies and researchers, as it serves as a strong foundation and background on biomarkers for complicated appendici- tis. Ultimately, this could lead to more optimal research that will benefit patients with complicated appendicitis, so they can be offered timely surgical intervention and thus improved treatment that will reduce the patient’s risk of postoperative complications.

Funding source

This research received funding from the Herlev Gentofte Hospital Research Council. The funding had no role in the study design; in the collection, analysis, and interpretation of data; in the writing of the report; and in the decision to submit the article for publication.

CRediT authorship contribution statement

Binyamin Sikander: Writing – original draft, Visualization, Project administration, Investigation, Funding acquisition, Formal analysis, Data curation, Conceptualization. Jacob Rosenberg: Writing – review & editing, Supervision, Resources, Project administration, Methodology, Conceptualization. Siv Fonnes: Writing – review & editing, Supervision, Project administration, Methodology, Conceptualization.

Declaration of Competing Interest

The authors declare that they have no competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

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