" we want to share with other interested colleagues,our adventure into surprises of rotator cuff tendinopathy; obviously a cooperative effort could give more consistent data " Manuel Branes

1. Shoulder Rotator Cuff Tendinopathy (SRCT).
M. Brañes A. (Orthopaedics). Adjunct Professor, Faculty of Sciences - University of Chile.
L. Arellano H., (Pathologist). Associated Professor, Faculty of Medicine - University of Chile.
J. Mackiness S. (Radiologist). Bio-Surgery Unit (ESWT) - AraucoSalud Clinic- Santiago, Chile.
"Until the biologist and the physical scientist speak the same language,
we will continue to struggle with trial-and-error solutions to healthcare problems ".
Frederick H. Silver [38].
Introduction.
Western societies are showing increasing numbers of patients suffering shoulder dis-
comfort and complains due to rotator cuff tendinopathy, pathology that encompasses
from symptomatic tendinopathy through massive rotator cuff tears. Surgical results for
rotator cuff tears repairs reported during last years are indicating the occurrence of
significant re-tear rates [1, 2]. This situation implies the necessity to focus attention on
the normal ontological repair mechanisms of this structure and re-orientated our efforts
towards better histological analysis, which could predict the healing condition of the
tissue and probabilities for re-tears.The aim of this review is to identify advances in
histopathology and characterizations of rotator cuff tendinopathy [3,4,5]. From our
clinical and histopathological experience, we formulate a correspondence for
histological and immuno-histochemical results, exploring possible links with
sonographic studies done upon a series of patients. For the reason that we use ESWT in
non-calcified Rotator Cuff Tendinopathy (single session 4000 pulses- 0,3mJ/mm2,
electro-hydraulic device (Orthospec/Medispec) or electro-magnetic device (Storz
Duolith)), our results, examples and commentaries will be added to the general
discussion.
Histopathological Classifications.
Recent reports describe different classifications methods using common histological
stains (Hematoxiline-Eosin and Alcian Blue stains), but at the moment to examine the
reliability of histopathological interpretation, done by experimented and interested
pathologists, the agreement is just defined as "acceptable" [6, 7]. In fact, it is difficult to
recognize specific patterns in this evolving tendinopathy and the scenario worse when a
limited number of histological techniques are applied. However, significant data are
obtained from published reports, for example, - rotator cuff tendinopathy is an
identifiable entity per se, - comparison of tendinopathic rotator cuffs permits to
ascertain a progression of histological damage, - tendons from healthy aged people
display minor histological differences compared with younger controls [3, 4, 6,7, 8, 10].
We use the classification proposed by Dr. G.P. Riley [8], which is based on the orga-
nization of tendon fiber bundles examined on hematoxylin-eosine stains, according a
useful four-point scale (Table 1). This arrangement emphasizes the importance of
structural integrity of the collagens in normal tendons and cell nuclei display, as well as
depicts their variations from standard disposition. From a mechanosensing standpoint,
adequate collagen array permits proper mechanotransduction; disruption of these
collagens rows integrity, induce different grades of upset on the way to transmit force
inside the tendon, situation that provoke dysfunction on cell-to-cell and matrix
components [9, 10, 38].

2. 2
In this point starts what it has been defined as "tendinopathy", a very complex cellular
& matrix (metabolic) process that encompasses variable degrees of acute as well as
chronic inflammation [11], both processes acting simultaneously over uneven
proportions of tendon tissues (Fig.1). We have branded two precise histological features
with sonographic expression, permitting to obtain an idea about the tendinopathic
matrix condition: the presence of micro-calcifications and/or intra-mural micro-tears.
Both instances were incorporated into Riley Classification (Table 1) and can be
followed by serial sonographic studies; we have seen their regression in tendinopathic
patients treated with ESWT in terms of 12 to 24 weeks after treatment, also in
accordance with improvements of clinical complains.
Table I. Riley Classification (modified).
Normal , Grade I Mild Moderate Severe
Degeneration , Grade II Degeneration , Grade III Degeneration , Grade IV
-elongated -shorter / still oval -increased in number -decreased number
-normal chromatin
Tenocyte -long axis parallel to -darker-staining chromatin -darker-staining chromatin -small / dark and rounded
Nuclei collagen bundles
-nuclei arranged in chains -loss of orientation
nuclei/ collagen fibers
-wavy outline - loosening of waviness -collagen hyalinization -diffuse collagen hyalinization
Tendon -individual fibers easily -initial disorientation of collagen -homogeneous eosinophilic -complete loss of orientation of
Fiber discernible in bundles fibers. staining collagens bundles.
Bundles
- hyper & hypo-echographic -initial chondroid metaplasia -advanced chondroid metaplasia
Histological signals
& -matrix micro-tears ( 1-3 mms) -micro-tears coalescence (> 4 mms)
Sonographic - non micro-tears
-"spotting" micro-calcifications -frank enthesis damage
Features - non micro-calcifications
added -calcifications (variable sizes) -complete rotator cuff tear
- normal enthesis
-initial enthesis damage
Obs: Histological findings of "micro-calcifications" tend to occurs in areas of initial collagen
hyalinization (corresponding to moderate degeneration, grade III); Calcifications of variable sizes are
referred to Calcified Tendinopathy. Intra-mural micro-tears in type III considered no longer than 3
mms, in type IV considered no longer than 7 mms.
Trying to solve limitations just using H-E and Alcian Blue stains, we expanded our
stains string including immuno-histochemical stains (PCNA, CD14, CD34, Tenascin-C,
Col I, Col III) and also concentrate our analysis upon the vascular component of
tendinopathy and their features, in order to obtain clues that could orientate to a repair
capability on this condition. A subset of tendinopathies (obtained from patients with
complete tears that received SW 8 – 10 weeks preoperatively) was also studied and their
findings incorporated. PCNA marker defines cells activated towards mitosis and
differentiation, CD14 and CD34 are used as markers for neo-endothelial cells (new cells
involved in neo-angiogenesis/vasculogenesis process, reflecting repair efforts) and
Tenascin-C is a glycoprotein expressed in areas of normal neo-angiogenesis/post natal
vasculogenesis and zones of tissue repair. Col I and Col III antibodies display
disposition for both collagens in altered tendons, also permitting to evaluate their
synthesis in areas of repair and remodeling. A very interesting work done by Dr. Hirose
[12] in rabbits, shown the utility of PCNA and Col I & III markers to evaluate the
tendon healing process during induced tear repair.

3. 3
Histopathological Findings.
To date our histological data bank comprises 65 cases studied; patient's average age is
62 years old (range: 48 to 74 y-o). Distribution into Riley's classification indicates that
5% of samples are type II, 64% correspond to type III and 31% fall into type IV
category. We obtain rotator cuff samples as a routine in open surgeries (consent
informed). Unfortunately those biopsies derived from arthroscopic procedures, contain
small amounts of tissue (+/- 3 x 3mms) and significant tissue percentage is disrupted by
the punch; in contrast, open surgery provides better and large samples in continuity,
facilitating the analysis of tissues abnormalities and their precise extensions. Each
sample is analyzed very carefully, taking micro-photographs (4x, 10x, 40x, 100x),
focused in matrix features, vascular supply and characteristics of its cellular
components; for automated images analysis, we send 5 microphotographs 40x for each
marker (IHC). Sections are examined according schedule, from bursa to articular areas
and from lateral to medial zones (fig.2). Semi-quantitative analysis (10x objectives) can
be done by using 10x10 grilles (100 chambers) on stained or well-formed structures.
In general, does not exist a direct relationship between the size of the rupture and degree
on histological abnormalities; small complete tears (less than 1 cm) can display deep
damage of tissue architecture in a diffuse form. Hashimoto [13] findings corroborate
this, but Matthews [14] have shown significant cellular changes as tear size increases,
influencing the healing rate. In fact, there are confounding factors when we try to find a
relationship among different variables as patient's age, duration of symptoms, tear size,
existence of common co-morbidities and possible histological patterns. We should
consider that repair capabilities came from matrix tendon circumstances related to
vascular supply responses and its cellular contributions; lasting results are expressed as
synthesis of neo-collagens and new cellular de-differentiated component, which replace
those lost by apoptosis or necrosis mechanisms.
Integrative data from histology and immunohistochemistry (that permits to evaluate the
possible healing potential of the tissue) with specific sonographic signal correlations,
may conduit to a better featuring of the tendinopathic tissue condition in our patients.
- 1.Vascular Responses and Blood Supply.
Normal human tendons exhibit a "vascular percentage area" on histological examination
equivalent to 2% to 3% and shoulder rotator cuff tendons accomplish this rule, but
vascular responses may modify this percentage (fig. 3). We have found in tendinopathic
tissue an average ”vascular percentage area” equivalent to 15%; after SW this number
arises up to 23% - 28% on tendon matrix able to bear this responses.
It is important to note that normal vascular repair mechanism display two inter-
connected instances defined as neo-angiogenesis and post-natal vasculogenesis; both
are joined steps acting in healing & remodeling for tendon tissues, as well as other
structures [15]. However, in chronic damage of tendons we have seen a gamut of neo-
angiogenesis foci, from “normal or reparative neo-angiogenesis” (fig.4), until frank
“abnormal neo-angiogenesis” (fig.7b, 7c, 8a, b, c, d).
For cases Riley type II, we have also found signs of vascular dilatation and initial neo-
angiogenesis from endotendon areas (fig.2b) as a common process.

4. 4
Fig.1. Diagram of four-levels of inflammation mechanisms (reproduced with permission (11)).
Fig.2a, left. Tendon structure and endotendon areas appears normal, corresponding to Riley type I.
Fig.2b, center. Initial neo-angiogenesis (white arrow) derived from endo-tendon area.
Fig.3, right. For comparison related to possible changes that this structure can display, a profuse neo-
vascularization with a calculated percentage of vascular area equivalent to 54%, is showed in case of
Shoulder Capsulitis (Frozen Shoulder stage).
Reparative neo-angiogenesis implicate normal blood vessel developed from a parent
one, displaying adequate number and disposition of pericytes along the new vessels,
with presence of red blood cells and fluid flow (active vessels) (fig.4 a).

5. 5
“Branching vessels” is an important aspect of normal fields that contains active neo-
angiogenesis (fig.4-b, c). It is possible to see hyper-cellularity in the vicinity of neo-
blood vessels, migrating from the lumen of vessels towards extra-cellular matrix (ECM)
(fig. 5a). These cells are polygonal in shape, many times with nucleoli visible in H-E
stains and afford in IHC techniques a positive PCNA markation indicating that are
activated for mitosis and differentiation, as well as CD133 positive markation (fig.5 b-c)
it is indicative that their origin is hematopoietic (Bone Marrow Stromal Cells, BMSC).
According recent reports, those considered BMSC CD133 cells might have other
origins as well [16]. New endothelial cells in neo-blood vessels express CD14 and
CD34 marker (fig.4-c), also indicative of their hematopoietic origin. This type of
reparative neo-angiogenesis does not display hemorrhagic foci and examined under
Alcian Blue stains, normally show well balanced expression of proteoglycans along
complete vascular structures (figs. 4-b). This reparative aspect can be seen in type II
tendinopathies, neighborhood to normal or not damaged tendon tissue; their frequency
in more advanced tendinopathies scarce progressively.
Fig. 4 a-left, normal neoangiogenesis (black arrows) from mature sub-synovial vessels (H-E, 10x). Fig. 4
b-center, image of normal neoangiogenesis depicting adequate disposition of proteoglycans (black
arrows) along active, well-covered pericytes vessels (Alcian Blue, 10x & zoom). Fig. 4 c-right, branching
in normal neo-angiogenesis (white arrow), depicted by CD34+ expression (IHC for CD34, 40x).
Fig. 5 a-left, hyper-cellularity, derived from neo-angiogenesis foci, invading tendon tissue (H-E, 10x). Fig
5 b-center, similar to fig.5-a, showing PCNA+ marked cells also moving away from vessels into tissue
(black arrows) (IHC for PCNA, 40x). Fig 5 c-right shows CD133 good markation in similar cells, indica-
tive of their hematopoietic origin. (IHC for CD133, 100x & zoom).

6. 6
On tendinopathic fields, we also found others two dissimilar vascular responses:
nodular and diffuse neo-angiogenesis foci, both types deserving a description of their
light microscope features.
Nodular neo-angiogenesis has been observed near to sub-synovial endotendon areas
and it is characterized by neo-blood vessel in clusters, well surrounded by pericytes,
plus high percentage of actives blood-vessels, containing red blood cells. It is observed
expression of neo-collagen synthesis and defined cellular component inside nodes; their
spontaneous frequency has been established close on 10 to 15% of cases from revised
material (fig 6). Their presence offers sometimes an acute contrast between its well-
organized architecture (node) and disorganized environmental tissue (matrix tendon). Its
position close to sub-synovial endotendon zones could be related to the caliber of blood
supply existing in these areas.
In comparison, diffuse neo-angiogenesis display an infiltrative mode in tendon tissue,
the coverage of pericytes is variable with micro-hemorrhagic foci associated to
undersupplied pericytes areas (fig.7, 8b, 8c). A frequent observation is that new vessels
are not grouped and significant numbers of them are inactive (does not contain red
blood cells), indicating that fluid flow is absent, meaning an unsuccessful neo-
angiogenesis [17]. In our vision, this is a capital sign of vascular disturbance,
developed on this pathology (as well as was pointed out by Nirschl et al., when
described similar histopathologic features in chronic epicondylitis). This ineffective
neo-angiogenesis tends to be hypo-cellular, not permitting an adequate trespassing of
activated cells towards tendon matrix.
Fig.6 a-left, spontaneous nodular neo-angiogenesis foci. Collagens of neo-synthesis display disorientation
and different pictorial properties (white arrows) in relation to tendon degenerate collagens (black arrows)
(H-E, 10x). Fig. 6 b-center, better characterization of neo-collagens disposition inside nodes (black
arrows), (Tri-chrome Masson stain, 10x). Fig 6 c-right, showing disposition of neo-collagens (black
arrows) surrounding node vascular component; notice that neo-vessels are well-covered by pericytes and
does not exist micro-hemorrhages (white arrows), (Toluidine Blue stain, 10x).
According tendinopathies transit from Riley III to Riley IV, the percentage vascular
area lessen and surrounding cellularity can be scanty in accordance to higher degree of
degeneration (fig.8), also expressing moderate to low stain-levels for CD34, CD14,
PCNA and Tenascin-C (these low markations are indicative of a weak potential for
healing, fig.9). Diminished vascular areas in type IV could be related to augmented
expression of chondral metaplasia observed in more degenerative tendon tissues, as
fragments of collagen IV and XVIII have anti-angiogenic activity [18, 19].

7. 7
Fig. 7 a-left, microphotograph displaying a quiescent vascular endotendon area in tendinopathy type II
(encircled dotted line) but in nearby areas it is appreciated a disperse neo-angiogenesis (white arrows),
(H-E,10x-zoom). Fig. 7-b, center: descendent neo-angiogenesis foci from sub-synovial endotendon areas
(dotted double arrow), displaying important micro-hemorrhagic areas along entire structure (black
arrows). Those hemorrhages are probably associated to instability of nascent neo-vessels because no
proper pericytes sheath (H-E, 10x). Fig.7 c-right, tendinopathy type II, showing infiltrative nature of
diffuse neo-angiogenesis, which include many inactive blood vessels, not well-covered by pericytes
(white arrows). (H-E, 10x & zoom).
Fig.8 a-left, type III tendinopathy showing hyper-cellularity in areas of infiltrative neo-angiogenesis, also
depicting hemorrhagic zones in tendon tissues (white arrows). Hemorrhagic areas associated to tendon
micro-tears are also visible (black arrows), (H-E,10x). Fig.8 b-center left, in contrast, hypo-cellular neo-
angiogenesis foci with marked absence of normal pericytes and inactive blood-vessels (black arrows),
(Alcian Blue stain, 40x). Fig 8 c-center right, showing vascular dilatation and significant scarcity of
pericytes coverage accompanied of hypo-cellularity (black arrows), (Masson Tri-chrome stain, 100x).
Fig.8 d-right, advanced tendinopathy type IV, depicting profound alterations in vascular architecture (no
presence of endothelial cells or pericytes), developed in areas of chondroid metaplasia.
Fig.9 a-left, PCNA+ expression in tendinopathy type III (black arrows), showing 60 nuclei which 25
display PCNA+ marker (41%) (40x). Fig.9 b-center left, Tenascin-C expression on same patient, shows a
weak and diffuse staining surrounding blood vessels areas (black arrows) (40x). Fig 9 c-center right,
adequate CD34 expression in endothelial cells of neo-blood vessels (40x). Fig.9 d-right, CD14 expression
in neo-vessels is feeble compared to CD34 (black arrows).

8. 8
According numbers available, in tendinopathies type III the average expression of PCNA
reach 35% of positive cells, most of them constitutive of blood vessels or in their
vicinity. Calcified tendinopathies showed a spontaneous up to 55% PCNA+ expression,
and this mark could be related to better tendon matrix quality in this specific
tendinopathy. In our study, Tenascin-C expression was in concordance with PCNA stain
intensity and always in vascular areas or close to them. CD34 and CD14 markers
display variable stain intensity, not in relation to observed PCNA-Tenascin-C
relationship. Also, mature endothelial cells of resident blood-vessels does not express
CD34 marker, its variable profile is indicative of neo-angio/vasculogenic process [39].
After SW treatments neo-angiogenesis adopt many times the nodular form, but
appears distributed randomly along treated matrix tendon; also we have seen PCNA+
expression equivalent to 80% to 85%, displayed on cells constitutive of neo-blood
vessels but also migrating into matrix. Tenascin-C expression is corresponding higher,
covering from new blood vessels to tendon tissue areas that underwent repair or reveals
zones of very initial vascular development. Neo-angiogenesis furthermore is augmented
and CD34 stain intensity is higher than those observed in tendinopathies non SW-
treated (fig.10). These SW-induced nodes express (beside new and active blood-vessels
& improved cellular environment with high PCNA+ index) distinct expression of
CD133, PDGF-bb and TGF-b1 (data not shown). A recent research reported by
Tempfer et al. [20], indicate that perivascular cells of the human supraspinatus tendon
display markers associated to tendon cells and stem-cells. Others investigations are
suggesting that pericytes have a role more definite in repair and remodeling process
because they are cells with demonstrated de-differentiation ability, so might help to
repopulate damaged areas and also are capable for synthesizing collagens[21, 22].
However, these significant improvements in markations are related only to type II and
III subsets of Riley's Classification, because in type IV is not possible to observe same
intensity of signals. Notice that type IV correspond to severe degeneration of the tissues
and in this situation occurs diffuse apoptosis with augmented chondroid metaplasia;
vascular supply is almost absent and its structure is severely anomalous (fig.8-d). Inside
our reviewed material that received SW pre-operatively, we have not seen a histological
response indicative of repair in those chondroid areas. For us, this finding is suggesting
that we should use SW in early stages of rotator cuff tendinopathies (type II - III),
following their clinical evolution with care. On the other hand, it is indicative of
performing an adequate trimming of the tear's margin during surgical repair, because
those chondroid areas occupy zones close to the tear's edge and neighborhood
fenestrated tissues. Matthews T. [23], reviewed histological features in a series of
patients with re-tears after surgical repair; his findings showed significant expression of
chondroid metaplasia areas in those patients, ranking 60% to 77%. In our vision,
chondroid degeneration of the tissue is the last common-path possible for this entity and
we think that is not able to repair in any way (figs.11 - 13 - 14).
- 2. Tendon Matrix Features. Collagens Analysis.
The techniques applied to collagen component are related to Col-I and Col-III
antibodies. It is know that the normal rate Col-I/Col-III for tendon tissues is 95:3 and
this figure is altered in tendinopathies, being lower level for Col-I and increasing levels
for Col-III [4].

9. 9
In the classification proposed it has been well described the morphological featuring of
collagens to light microscope; our studies using antibodies for both collagens, suggest
that structure and quality tendon are affected in a similar way as Classification advice
(Fig.12, Fig.13). According these examples showed, is understandable that we obtain
better results with ESWT in cases of tendinopathies type II and III, where tendon matrix
Fig.10 a-left, PCNA expression in tendinopathy type III SW-treated 8 weeks earlier.71 nuclei seen, 79%
PCNA(+) (white arrows) many of them migrating into matrix tendon, 21% PCNA(-) (black arrows).
Fig.10 b-center left, Tenascin-C distribution around blood-vessels and also beyond that margin (white
arrows). Fig.10 c-center right, noteworthy CD34 expression. Fig.10 d-right, positive CD14 markation.
All figures 40x, same patient.
Fig.11 a-left, PCNA expression in tendinopathy type IV SW-treated 8 weeks earlier. Just 11 nuclei seen
in cells resembling chondrocytes (acellular-zone); their PCNA(+) expression possibly is also related to
their intrinsic metabolic rate & micro-environment plus SW (chondroid metaplasia area). Fig.11 b-center
left, Tenascin-C markation is absent, even when some abnormal vascular areas are displayed in the
picture (black arrows); the entire sequence in search of the Tenascin-C spot in this patient was negative.
Fig.11 c-center right, CD34 expression, showing very weak spots (black arrows). Fig.11 d-right, CD14
expression, also displays a feeble stain. All figures 40x, same patient.
appears more normal and able to bear a new reparative neo-angiogenesis/vasculogenesis
process (Figs. 10 - 15). The metabolic condition of the matrix it seems to be extreme in
importance, the cellular component (and its behavior) is reflecting matrix status and also
it is known that normal mechanotransduction maintain a proper balance of matrix
metallo-proteases (MMP's), one of the critical steps in matrix responses. Recent
published works are indicating that SW has also the capacity to influence some
metabolic mechanisms in matrix tissues through molecular induction: nitric oxide (NO)
synthesis through (i)NOS is an example [24]. Also, in wounded skin (burns) it has been
demonstrated a modulating action upon cytokines release, lessening their acute
inflammatory component [25]. Further evidence was given by Aicher et al [26], when
demonstrated a higher incorporation of infused endothelial cells to ischemic tissue after
SW use (displaying an up-regulation for VEGF and SDF-1), in comparison when SW-
treatment was not applied just 24 hours earlier.

10. 10
According our literature reviews, cytokine SDF-1 and its receptor CXCR4 are able to
mediate endothelial progenitor cell (EPC's) homing [26, 27] on ischemic sites, and are
significantly up-regulated after 24 hours of SW-applied treatment [26] with subsequent
augmented incorporation of EPC's and resulting in neoangio/vasculogenesis repair
mechanisms. All these examples are suggesting that SW, beside its action on tissue
cells, might have an immediate and direct effect upon active or inactive ("sequestered")
molecular mediators resident in the matrix, corresponding to activity at “molecular
level” of inflammation diagram (fig.1).
Fig.12-left, tendinopathy type III, showing initial collagen hyalinization giving a "mottled appearance"
(H-E, 10x). Fig.12-center: Col I expression, displayed surrounding vessels, (40x). Fig.12-right, Col III
expression (intertwined with Col I) over equal area (sequential sections) (40x, same patient).
Fig. 13-left, tendinopathy type IV, depicting significant chondroid areas (black arrows) and a diffuse
pattern of collagen hyalinization surrounding them. Nuclei are small, dark and rounded (H-E, 10x).
Fig.13-center, Col I stain shows an abnormal pattern, suggesting greater compromise of tendon structure
(40x). Fig.13-right, Col III expression (same patient), corroborates impression about compromise of
tendon collagens because its appearance is disorganized and weak (40x).
Fig.14-left, tendinopathy type IV in a patient that received SW treatment, displaying matrix alterations
according categorization (H-E, 10x). Fig.14-center, Col I is abnormal (40x). Fig.14-right, Col III marker
outlook also indicates its anomaly (40x, same patient).
- 3. Microcalcifications.
Riley reported in 1996 his research about "dystrophic calcification" observed in cases of
tendinopathic rotator cuffs [28], also comparing differences with calcified tendinopathy;
other important contribution was completed by Archer in 1993, where describe matrix

11. 11
changes associated with calcifications in rotator cuffs, also including EM images of
different cellular types found in these conditions [29].
Fig.15-left, tendinopathy type III in a patient that received SW treatment. The figure display nodular neo-
angiogenesis foci (white dotted line) found in deep tendon, characterized by high number of active new
vessels in cluster with proper pericytes envelope, significant hyper-cellularity surrounding vascular
structures and no signs of hemorrhage. White rectangle emphasized an area in which tendon collagens
appears curved because the presence and activity of the node (suggesting that node expands itself). White
arrow indicates the axis of tendon native collagens (H-E, 10x). Fig.15-center shows neo-vascular areas,
inside the node, which display significant Col I synthesis (black arrows). Demarcating the border of the
node (white arrow & dotted line drawn), permits to obtain an idea of the intensity of Col I neo-synthesis
inside node. Fig.15-right, augmented Col III synthesis (inside node) and in relation to SW-neo-vascular
structures induced (black arrows) (40x, same patient).
The appearance of micro-calcifications (Fig.16) in tendon tissue is indicative of initial
hyalinization of the structure. It is possible to observe them contiguous to the frayed
border in case of tears, but most times are incorporated to tissue and not induces an
inflammatory response. Its sonographic incidence in symptomatic tendinopathies
reaches 30% to 40% and comprises small calcium deposits (2 to 4 mms); sometimes,
that deposit enlarges, giving the impression to correspond to Calcified Tendinopathy.
After SW treatments those micro-calcifications can disappear and its place is occupied
by almost normal tendon signals in echographic studies, suggesting that a remodeling
process occurred. This instance has been observed frequently and it is considered by us,
as an indication of SW-induced recovery of the tissue (fig.17).
Fig.16 left, "spotting" of micro-calcifications with hyalinization of the surrounding tissues (H-E, 10x &
zoom). Fig.16 center, sub-synovial disposition of micro-calcifications, surrounded by few cells in similar
ground (H-E, 10x). Fig.16 right, pretty picture obtained with 100x objective, demonstrating a single
micro-calcification surrounded by 12 cells (not fibroblasts), probably at early stage of resolution. Tendon
matrix put on view hyalinization of the ground (H-E, 100x).

12. 12
Fig.17. Micro-Calcification disappeared after SW treatment (single session). Its distribution involve most,
of the time, subscapularis and supraspinatus tendon, but tendinopathic aspect overwhelming the general
picture in the sonogram. However, a careful exam can offer some clues about what is occurring inside the
tendon.
- 4. Intra-mural Micro-Tears.
The appearance of micro-tears is another aspect in the natural history of tendinopathies,
caused initially by non-adequate balance in metabolic pathways: overuses stress those
debilitated structures causing intra-mural tearing. In clinical setting, patient's complaint
refers as a dull pain in the shoulder and many of them are displaying evolving intra-
mural tears on sonographic studies; on serial exams, we might see that micro-tears
grow-up (coalescence) and the patient's condition worsening progressively. Quite the
opposite, when we applied a SW treatment (single session, 4000 impulses,
0,30mJ/mm2) is possible to assist to improvement of patient's complaint and to see the
resolution of intra-mural tears in echographic studies 12 to 24 weeks later.
Fig.18, left. Histological section depicting an intra-mural tear (encircled dotted area) in which its seen a
stump of rotator cuff tissue (double white arrow, on left) recently detached from where an acute
hemorrhage its seen (double white arrow, on right); tendon cells in the stump will undergo apoptosis due
to absence of normal mechanotransduction. Black arrow shows normal tendon structure and white dotted
arrow display probable ischemic tendon tissue (pictorial upset). Fig.18, right. Echographic finding of
intra-mural single tear (4mms) in a patient with symptomatic tendinopathy; we have seen complete
restoration of tendon structure after SW treatment on similar cases.
We have defined an extension of 1 to 3 mms for type III and 4 to 7 mms in type IV for
these micro-tears found in matrix tendon; the upper limit suggested is in relation to

13. 13
anatomical measurements of the structure: a rupture of 8 mms to 10mm or higher, its
more probable to correspond to a complete rotator cuff tear (fig.19). Echographic
studies should be done patiently and watchfully on these populations selected for SW,
in order to get precise diagnosis for patient's condition an appropriate result using the
new technology. Is not uncommon to receive from the Echographist a report indicating
a “coarse ecographic signal” from patients that received SW treatment 18 or 24 weeks
earlier; asked him explains that his impression correspond to occurrence of a “ subtle
diffuse scarring “, and probably is indicating higher collagen synthesis in those treated
patients classified in grade II or III (fig.15).
.
Fig.19. Patient female, 48 years-old, medical assistant, 2 last years with progressive pain and shoulder
discomfort. On left, an intra-mural tear, clearly visible (white arrows). She received single SW treatment.
Echographic control (right) indicate healing in progress on selected area [13 weeks later], and also her
condition was improving. SW acts, in our opinion, as a "biological inducer of ontological repair
mechanisms", so healing time for tendon structures should be considered at least 12 weeks and multiple
of this (24weeks).
In summary, the classification of Dr. Riley permits to us to identify the condition of
tendinopathic matrix and immuno-histochemical analysis support its singular class long-
established principally by Col I and Col III performance. It is possible to evaluate, in a
better way, the healing probabilities using markers as PCNA-CD14-CD34-Tenascin-C,
Col I & III, comparing for their activity in tendon tissue as well as in vascular reactive
areas [30, 31]. We have found a particular vascular arrangement (nodular neo-
angio/vasculogenesis) in tendinopathies over sub-synovial areas; in SW-treated tendon
we found improved angiogenesis with significant expression of nodular forms, disposed
along the entire tendon structure, which include active hyper-cellularity. Our hypothesis
is that those nodular forms are in correspondence to post-natal vasculogenesis
mechanism described by Asahara [15], matching to a more active angiogenesis with
high population of de-differentiated cells, committed to replace lost cellular mass during
the entire process of tendinopathy and capable to initiate a remodeling activity of the
damaged matrix.
Kovacevic et al. [36] suggested that a normal insertion (enthesis zone) does not
regenerate after repair probably like to "abnormal or insufficient gene expression and/or
cell differentiation at the repair site". Wang et al [33] using SW as treatment in delayed
bone-tendon healing demonstrated improved regeneration of fibro-cartilage zone and
osteogenesis; concomitantly, very similar results were reported later by Qin [37].
From our experience, with SW clinical applications on different pathologies, we have

14. 14
been observing a "scarless healing" or a "supplementary reconstructive tissue repair",
meaning that this induced healing it is related with higher stem cells engraftment and/or
higher "stemness expression" from cellular component integrated for healing. This
observation is not new at the core of ISMST, but according passing time, more well-
built data from different sources support it [32-35, 37].
Clinical improvements of non-calcified shoulder tendinopathic conditions after SW
treatments, strongly suggest to us, that this new technology has a definite therapeutic
role on this specific set of human pathologies and deserve more cooperative research
efforts.
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